CA2416414A1 - Human kinases - Google Patents

Abstract

The invention provides human human kinases (PKIN) and polynucleotides which identify and encode PKIN. The invention also provides expression vectors, ho st cells, antibodies, agonists, and antagonists. The invention also provides methods for diagnosing, treating, or preventing disorders associatedd withd abberant expression of PKIN.

Description

HUMAN KINASES
TECHNICAL FIELD
This invention relates to nucleic acid and amino acid sequences of human kinases and to the use of these sequences in the diagnosis, treatment, and prevention of cancer, immune disorders, disorders affecting growth and development, cardiovascular diseases, and lipid disorders, and in the assessment of the effects of exogenous compounds on the expression of nucleic acid and amino acid sequences of human kinases.
l0 BACKGROUND OF THE INVENTION
Kinases comprise the largest known enzyme superfamily and vary widely in their target molecules. Kinases catalyze the transfer of high energy phosphate groups from a phosphate donor to a phosphate acceptor. Nucleotides usually serve as the phosphate donor in these reactions, with most kinases utilizing adenosine triphosphate (ATP). The phosphate acceptor can be any of a variety of molecules, including nucleosides, nucleotides, lipids, carbohydrates, and proteins. Proteins are phosphorylated on hydroxyamino acids. Addition of a phosphate group alters the local charge on the acceptor molecule, causing internal conformational changes and potentially influencing intermolecular contacts. Reversible protein phosphorylation is the primary method for regulating protein activity in eukaryotic cells. In general, proteins are activated by phosphorylation in response to extracellular signals such as hormones, neurotransmitters, and growth and differentiation factors. The activated proteins initiate the cell's intracellular response by way of intracellular signaling pathways and second messenger molecules such as cyclic nucleotides, calcium-calinodulin, inositol, and various mitogens, that regulate protein phosphorylation.
Kinases are involved in all aspects of a cell's function, from basic metabolic processes, such as glycolysis, to cell-cycle regulation, differentiation, and communication with the extracellular environment through signal transduction cascades. Inappropriate phosphorylation of proteins in cells has been linked to changes in cell cycle progression and cell differentiation.
Changes in the cell cycle have been linked to induction of apoptosis or cancer. Changes in cell differentiation have been linked to diseases and disorders of the reproductive system, immune system, and skeletal muscle.
There are two classes of protein kinases. One class, protein tyrosine kinases (PTKs), phosphorylates tyrosine residues, and the other class, protein serine/threonine kinases (STKs), phosphorylates serine and threonine residues. Some PTKs and STKs possess structural characteristics of both families and have dual specificity for both tyrosine and serine/threonine residues. Almost all kinases contain a conserved 250-300 amino acid catalytic domain containing specific residues and sequence motifs characteristic of the kinase family. The protein kinase catalytic domain can be further divided into 11 subdomains. N-terminal subdomains I-IV
fold into a two-lobed structure which binds and orients the ATP donor molecule, and subdomain V
spans the two lobes. C-terminal subdomains VI-XI bind the protein substrate and transfer the gamma phosphate from ATP to the hydroxyl group of a tyrosine, serine, or threonine residue. Each of the 11 subdomains contains specific catalytic residues or amino acid motifs characteristic of that subdomain. For example, subdomain I contains an ~-amino acid glycine-rich ATP binding consensus motif, subdomain II
contains a critical lysine residue required for maximal catalytic activity, and subdomains VI through IX
comprise the highly conserved catalytic core. PTKs and STKs also contain distinct sequence motifs in subdomains VI and VIII which may confer hydroxyamino acid specificity.
In addition, kinases may also be classified by additional amino acid sequences, generally between 5 and 100 residues, which either flank or occur within the kinase domain. These additional amino acid sequences regulate kinase activity and determine substrate specificity. (Reviewed in Hardie, G. and S. Hanks (1995) The Protein Kinase Facts Book, Vol I, pp. 17-20 Academic Press, San Diego CA.). In particular, two protein kinase signature sequences have been identified in the kinase domain, the first containing an active site lysine residue involved in ATP binding, and the second containing an aspartate residue important for catalytic activity. If a protein analyzed includes the two protein kinase signatures, the probability of that protein, being a protein kinase is close to 100%
(PROSITE: PDOC00100, November 1995).
Protein Tyrosine Kinases Protein tyrosine kinases (PTKs) may be classified as either transmembrane, receptor PTKs or nontransmembrane, nonreceptor PTK proteins. Transmembrane tyrosine kinases function as receptors for most growth factors. Growth factors~bind to the receptor tyrosine kinase (RTK), which causes the receptor to phosphorylate itself (autophosphorylation) and specific intracellular second messenger proteins. Growth factors (GF) that associate with receptor PTKs include epidermal GF, platelet-derived GF, fibroblast GF, hepatocyte GF, insulin and insulin-like GFs, nerve GF, vascular endothelial GF, and macrophage colony stimulating factor.
Nontransmembrane, nonreceptor PTKs lack transmembrane regions and, instead, form signaling complexes with the cytosolic domains of plasma membrane receptors.
Receptors that function through non-receptor PTKs include those for cytokines and hormones (growth hormone and prolactin), and antigen-specific receptors on T and B lymphocytes.
Many PTKs were first identified as oncogene products in cancer cells in which PTK

activation was no longer subject to normal cellular controls. In fact, about one third of the known oncogenes encode PTKs. Furthermore, cellular transformation (oncogenesis) is often accompanied by increased tyrosine phosphorylation activity (Charbonneau, H. and N.K. Tonks (1992) Annu. Rev.
Cell Biol. 8:463-493). Regulation of PTK activity may therefore be an important strategy in controlling some types of cancer.
Substrates for tyrosine kinases can be identified using anti-phosphotyrosine antibodies to screen tyrosine-phosphorylated cDNA expression libraries. Fish, so named for tyrosine-phosphorylated in Src-transfromed fibroblast, is a tyrosine kinase substrate which has been identified by such a technique. Fish has five SH3 domains and a phox homology (PX) domain. Fish is l0 suggested to be involved in signalling by tyrosine kinases and have a role in the actin cytoskeleton (Lock,P. et al (1998) EMBO J. 17:4346-4357).
SHP-2, an SH2-domain-containing phosphotyrosine phosphatase, is a positive signal transducer for several receptor tyrosine kinases (RTKs) and cytokine receptors. Phosphotyrosine phosphatases are critical positive and negative regulators in the intraellular signalling pathways that result in growth-factor-specific cell responses such as mitosis, migration, differentiation, transformation, survival or death. Signal-regulatory proteins (SIRPs) comprise a new gene family of at least 15 members, consisting of two subtypes distinguished by the presence or absence of a cytoplasmic SHP-2-binding domain. The SIRP-alpha subfamily members have a cytoplasmic SHP2-binding domain and includes SIRP-alpha-1, a transmembrane protein, a substrate of activated RTKs and which binds to SH2 domains. SIRPs have a high degree of homology with immune antigen recognition molecules. The SIRP-beta subfamily lacks the cytoplasmic tail. The SIRP-beta-1 gene encodes a polypeptide of 398 amino acids. SIRP family members axe generally involved in regulation of signals which define differnet physiological and pathological process (Kharitonenkov,A. et al (1997) Nature 386:181-186). Two possible areas of regulation include determination of brain diversity and genetic individuality (Sano,S et al (1999) Biochem. J. 344 Pt 3:667-675) and recognition of self which fails in diseases such as hemolytic anemia (Oldenborg,P.-A et al (2000) Science 288:2051-2054).
Protein SerinefThreonine Kinases Protein serine/threonine kinases (STKs) are nontransmembrane proteins. A
subclass of STKs are known as ERKs (extracellular signal regulated kinases) or MAPs (mitogen-activated protein kinases) and are activated after cell stimulation by a variety of hormones and growth factors.
Cell stimulation induces a signaling cascade leading to phosphorylation of MEK
(MAP/ERK kinase) which, in turn, activates ERK via serine and threonine phosphorylation. A
varied number of proteins represent the downstream effectors for the active ERK and implicate it in the control of cell proliferation and differentiation, as well as regulation of the cytoskeleton.
Activation of ERK is normally transient, and cells possess dual specificity phosphatases that are responsible for its down-regulation. Also, numerous studies have shown that elevated ERK activity is associated with some cancers. Other STKs include the second messenger dependent protein kinases such as the cyclic-AMP dependent protein kinases (PKA), calcium-calinodulin (CaM) dependent protein kinases, and the mitogen-activated protein kinases (MAP); the cyclin-dependent protein kinases; checkpoint and cell cycle kinases; Numb-associated kinase (Nak); human Fused (hFu);
proliferation-related kinases; 5'-AMP-activated protein kinases; and kinases involved in apoptosis.
The second messenger dependent protein kinases primarily mediate the effects of second l0 messengers such as cyclic AMP (cAMP), cyclic GMP, inositol triphosphate, phosphatidylinositol, 3,4,5-triphosphate, cyclic ADP ribose, arachidonic acid, diacylglycerol and calcium-calinodulin. The PKAs are involved in mediating hormone-induced cellular responses and are activated by cAMP
produced within the cell in response to hormone stimulation. cAMP is an intracellular mediator of hormone action in all animal cells that have been studied. Hormone-induced cellular responses include thyroid hormone secretion, cortisol secretion, progesterone secretion, glycogen breakdown, bone resorption, and regulation of heart rate and force of heart muscle contraction. PKA is found in all animal cells and is thought to account for the effects of cAMP in most of these cells. Altered PKA
expression is implicated in a variety of disorders and diseases including cancer, thyroid disorders, diabetes, atherosclerosis, and cardiovascular disease (Isselbacher, K.J. et al. (1994) Harrison's Principles of Internal Medicine, McGraw-Hill, New York NY, pp. 416-431, 1887).
The casein kinase I (CKI) gene family is another subfamily of serine/threonine protein kinases. This continuously expanding group of kinases have been implicated in the regulation of numerous cytoplasmic and nuclear processes, including cell metabolism, and DNA
replication and repair. CKI enzymes are present in the membranes, nucleus, cytoplasm and cytoskeleton of eukaryotic cells, and on the mitotic spindles of mammalian cells (Fish, K.J.
et al. (1995) J. Biol. Chem.
270:14875-14883).
The CKI family members all have a short amino-terminal domain of 9-76 amino acids, a highly conserved kinase domain of 284 amino acids, and a variable carboxyl-terminal domain that ranges from 24 to over 200 amino acids in length (Cegielska, A. et al. (1998) J.
Biol. Chem. 273:1357-1364).
The CKI family is comprised of highly related proteins, as seen by the identification of isoforms of casein kinase I from a variety of sources. There are at least five mammalian isoforms, a, (3, y, 8, and E. Fish et aL, identified CKI-epsilon from a human placenta cDNA library. It is a basic protein of 416 amino acids and is closest to CKI-delta. Through recombinant expression, it was determined to phosphorylate known CKI substrates and was inhibited by the CKI-specific inhibitor CKI-7. The human gene for CKI-epsilon was able to rescue yeast with a slow-growth phenotype caused by deletion of the yeast CKI locus, HRR250 (Fish et al., supra).
The mammalian circadian mutation tau was found to be a semidominant autosomal allele of CKI-epsilon that markedly shortens period length of circadian rhythms in Syrian hamsters. The tau locus is encoded by casein kinase I-epsilon, which is also a homolog of the Drosophila circadian gene double-time. Studies of both the wildtype and tau mutant CKI-epsilon enzyme indicated that the mutant enzyme has a noticeable reduction in the maximum velocity and autophosphorylation state.
Further, in vitro, CKI-epsilon is able to interact with mammalian PERIOD
proteins, while the mutant enzyme is deficient in its ability to phosphorylate PERIOD. Lowrey et al., have proposed that CKI-epsilon plays a major role in delaying the negative feedback signal within the transcription-translation-based autoregulatory loop that composes the core of the circadian mechanism.
Therefore the CKI-epsilon enzyme is an ideal target for pharmaceutical compounds influencing circadian rhythms, jet-lag and sleep, in addition to other physiologic and metabolic processes under circadian regulation (Lowrey, P.L. et al. (2000) Science 288:483-491).
Homeodomain-interacting protein kinases (HIPKs) are serinelthreonine kinases and novel members of the DYRK kinase subfamily (Hofmann, T.G. et al. (2000) Biochimie 82:1123-1127).
HIPKs contain a conserved protein kinase domain separated from a domain that interacts with homeoproteins. HIPKs are nuclear kinases, and HIPK2 is highly expressed in neuronal tissue (Kim, Y.H. et al. (1998) J. Biol. Chem. 273:25875-25879; Wang, Y. et al. (2001) Biochim. Biophys. Acta 1518:168-172). HIPKs act as corepressors for homeodomian transcription factors. This corepressor activity is seen in posttranslational modifications such as ubiquitination and phosphorylation, each of which are important in the regulation of cellular protein function (Kim, Y.H.
et al. (1999) Proc. Natl.
Acad. Sci. USA 96:12350-12355).
The marine homology to Caenorhabditis elegans UNC51, a serinelthreonine kinase, has been determined to be required to signal the program of gene expression leading to axon formation from granule cells of the cerebellar cortex (Tomoda, T. et al (1999) Neuron 24:833-346. The human homolog of UNC-51, ULK1, for UNC-51 (C. elegans)-like kinase 1, is composed of 1050 amino acids, has a calculated MV of 112.6 kDa and a pI of 8.80. ULK1 has 41 % overall sequence similarity to UNC-51 and is highly convserved among vertebrates including mammals, birds, reptiles, amphibians, and fish. By Northern blot analysis, Kuroyanagi et al have shown ULK1 to be ubiquitously expressed in adult tissues, including skeletal muscle, heart, pancreas, brain, placenta, liver, kidney, and lung while UNC-51 has been specifically located in the nervous system of C. elegans. Fish and RH mapping confirmed the localization of ULKl to human chromosome 12q24.3. (Kuroyanagi, H. et al (1998) Genomics 51:76-85.
Calcium-Calmodulin Dependent Protein Kinases Calcium-calmodulin dependent (CaM) kinases are involved in regulation of smooth muscle contraction, glycogen breakdown (phosphorylase kinase), and neurotransmission (CaM kinase I and CaM kinase II). CaM dependent protein kinases are activated by calmodulin, an intracellular calcium receptor, in response to the concentration of free calcium in the cell. Many CaM kinases are also activated by phosphorylation. Some CaM kinases are also activated by autophosphorylation or by other regulatory kinases. CaM kinase I phosphorylates a variety of substrates including the l0 neurotransmitter-related proteins synapsin I and II, the gene transcription regulator, CREB, and the cystic fibrosis conductance regulator protein, CFTR (Haribabu, B. et al.
(1995) EMBO J. 14:3679-3686). CaM kinase II also phosphorylates synapsin at different sites and controls the synthesis of catecholamines in the brain through phosphorylation and activation of tyrosine hydroxylase. CaM
kinase II controls the synthesis of catecholamines and seratonin, through phosphorylation/activation of tyrosine hydroxylase and tryptophan hydroxylase, respectively (Fujisawa, H.
(1990) BioEssays 12:27-29). The mRNA encoding a calmodulin-binding protein kinase-like protein was found to be enriched in mammalian forebrain. This protein is associated with vesicles in both axons and dendrites and accumulates largely postnatally. The amino acid sequence of this protein is similar to CaM-dependent STKs, and the protein binds calmodulin in the presence of calcium (Godbout, M.
et al. (1994) J.
Neurosci.l4:1-13).
Mito~en-Activated Protein Kinases The mitogen-activated protein kinases (MAP) which mediate signal transduction from the cell surface to the nucleus via phosphorylation cascades are another STK family that regulates intracellular signaling pathways. Several subgroups have been identified, and each manifests different substrate specificities and responds to distinct extracellular stimuli (Egan, S.E. and R.A. Weinberg (1993) Nature 365:781-783). There are 3-kinase modules comprising the MAP
kinase cascade:
MAPK (MAP), MAPK kinase (MAP2K, MAPKK, or MKK), and MKK kinase (MAP3K, MAPKKK, OR MEKK) (Wang,X.S. et al (1998) Biochem. Biophys. Res. Commun. 253:33-37). The extracellular-regulated kinase (ERK) pathway is activated by growth factors and mitogens, for example, epidermal growth factor (EGF), ultraviolet light, hyperosmolar medium, heat shock, endotoxic lipopolysaccharide (LPS). The closely related though distinct parallel pathways, the c-Jun N-terminal kinase (JNK), or stress-activated kinase (SAPK) pathway, and the p38 kinase pathway are activated by stress stimuli and proinflammatory cytokines such as tumor necrosis factor (TNF) and interleukin-1 (IL-1). Altered MAP kinase expression is implicated in a variety of disease conditions including cancer, inflammation, immune disorders, and disorders affecting growth and development.. MAP
kinase signaling pathways are present in mammalian cells as well as in yeast.
MAPKKK6 (MAP3K6) is one of numerous MAP3Ks identified. Isolated from skeletal muscle, MAP3K6 is 1,280 amino acids in length with 11 kinase subdomains and is detected in several tissues. The highest expression has been found in heart and skeletal muscle.
MAP3K6 has 45%
amino acid sequence identity with MAP3K5, while their catalytic domains share 82%o identity.
MAP3K6 interaction with MAP3K5 ira vivo was confirmed by coimmunoprecipitation. Recombinant MAP3K6 has been shown to weakly activate the JNK but not the p38 kinase or ERK
pathways (Wang,X.S. et al. supra) Cvclin-Dependent Protein Kinases The cyclin-dependent protein kinases (CDKs) are STKs that control the progression of cells through the cell cycle. The entry and exit of a cell from mitosis are regulated by the synthesis and destruction of a family of activating proteins called cyclins. Cyclins are small regulatory proteins that bind to and activate CDKs, which then phosphorylate and activate selected proteins involved in the mitotic process. CDKs are unique in that they require multiple inputs to become activated. In addition to cyclin binding, CDK activation requires the phosphorylation of a specific threonine residue and the dephosphorylation of a specific tyrosine residue on the CDK.
Another family of STKs associated with the cell cycle are the NIMA (never in mitosis)-related kinases (Neks). Both CDKs and Neks are involved in duplication, maturation, and separation of the microtubule organizing center, the centrosome, in animal cells (Fry, A.M. et al. (1998) EMBO J.
17:470-481 ).
Checkpoint and Cell Cycle Kinases In the process of cell division, the order and timing of cell cycle transitions are under control of cell cycle checkpoints, which ensure that critical events such as DNA
replication and chromosome segregation are carried out with precision. If DNA is damaged, e.g. by radiation, a checkpoint pathway is activated that arrests the cell cycle to provide time for repair.
If the damage is extensive, apoptosis is induced. In the absence of such checkpoints, the damaged DNA is inherited by aberrant cells which may cause proliferative disorders such as cancer. Protein kinases play an important role in this process. For example, a specific kinase, checkpoint kinase 1 (Chkl), has been identified in yeast and mammals, and is activated by DNA damage in yeast. Activation of Chk1 leads to the arrest of the cell at the G2/M transition (Sanchez, Y. et al. (1997) Science 277:1497-1501). Specifically, Chk1 phosphorylates the cell division cycle phosphatase CDC25, inhibiting its normal function which is to dephosphorylate and activate the cyclin-dependent kinase Cdc2. Cdc2 activation controls the entry of cells into mitosis (Peng, C.-Y. et al. (1997) Science 277:1501-1505). Thus, activation of Chkl prevents the damaged cell from entering mitosis. A similar deficiency in a checkpoint kinase, such as Chkl, may also contribute to cancer by failure to arrest cells with damaged DNA at other checkpoints such as G2/M.
Proliferation-Related Kinases Proliferation-related kinase is a serum/cytokine inducible STK that is involved in regulation of the cell cycle and cell proliferation in human megakarocytic cells (Li, B. et al. (1996) J. Biol. Chem.
271:19402-19408). Proliferation-related kinase is related to the polo (derived from Drosophila polo gene) family of STKs implicated in cell division. Proliferation-related kinase is downregulated in lung tumor tissue and may be a proto-oncogene whose deregulated expression in normal tissue leads~to oncogenic transformation.
5'-AMP-activated protein kinase A ligand-activated STK protein kinase is 5'-AMP-activated protein kinase (AMPK) (Gao, G.
et al. (1996) J. Biol Chem. 271:8675-8681). Mammalian AMPK is a regulator of fatty acid and sterol synthesis through phosphorylation of the enzymes acetyl-CoA carboxylase and hydroxymethylglutaryl-CoA reductase and mediates responses of these pathways to cellular stresses such as heat shock and depletion of glucose and ATP. AMPK is a heterotrimeric complex comprised of a catalytic alpha subunit and two non-catalytic beta and gamma subunits that are believed to regulate the activity of the alpha subunit. Subunits of AMPK have a much wider distribution in non-lipogenic tissues such as brain, heart, spleen, and lung than expected.
This distribution suggests that its role may extend beyond regulation of lipid metabolism alone.
Kinases in Apoptosis Apoptosis is a highly regulated signaling pathway leading to cell death that plays a crucial role in tissue development and homeostasis. Deregulation of this process is associated with the pathogenesis of a number of diseases including autoimmune disease, neurodegenerative disorders, and cancer. Various STKs play key roles in this process. ZIP kinase is an STK
containing a C-terminal leucine zipper domain in addition to its N-terminal protein kinase domain.
This C-terminal domain appears to mediate homodimerization and activation of the kinase as well as interactions with transcription factors such as activating transcription factor, ATF4, a member of the cyclic-AMP
responsive element binding protein (ATF/CREB) family of transcriptional factors (Sanjo, H, et al.
(1998) J. Biol. Chem. 273:29066-29071). DRAK1 and DRAK2 are STKs that share homology with the death-associated protein kinases (DAP kinases), known to function in interferon-'y induced apoptosis (Sanjo et al., supra). Like ZIP kinase, DAP kinases contain a C-terminal protein-protein interaction domain, in the form of ankyrin repeats, in addition to the N-terminal kinase domain. ZIP, DAP, and DRAK kinases induce morphological changes associated with apoptosis when transfected into NIH3T3 cells (Sanjo et al., supra). However, deletion of either the N-terminal kinase catalytic domain or the C-terminal domain of these proteins abolishes apoptosis activity, indicating that in addition to the kinase activity, activity in the C-terminal domain is also necessary for apoptosis, possibly as an interacting domain with a regulator or a specifc substrate.
RICK is another STK recently identified as mediating a specific apoptotic pathway involving the death receptor, CD95 (Inohara, N. et al. (1998) J. Biol. Chem. 273:12296-12300). CD95 is a member of the tumor necrosis factor receptor superfamily and plays a critical role in the regulation and homeostasis of the immune system (Nagata, S. (1997) Cell 88:355-365). The CD95 receptor signaling pathway involves recruitment of various intracellular molecules to a receptor complex following ligand binding. This process includes recruitment of the cysteine protease caspase-8 which, in turn', activates a caspase cascade leading to cell death. RICK is composed of an N-terminal kinase catalytic domain and a C-terminal "caspase-recruitment" domain that interacts with caspase-like domains, indicating that RICK plays a role in the recruitment of caspase-8.
This interpretation is supported by the fact that the expression of RICK in human 293T cells promotes activation of caspase-8 and potentiates the induction of apoptosis by various proteins involved in the CD95 apoptosis pathway (Inohara et al., supra).
Mitochondria) Protein Kinases A novel class of eukaryotic kinases, related by sequence to prokaryotic histidine protein kinases, are the mitochondria) protein kinases (MPKs) which seem to have no sequence similarity with other eukaryotic protein kinases. These protein kinases are located exclusively in the mitochondria) matrix space and may have evolved from genes originally present in respiration-dependent bacteria which were endocytosed by primitive eukaryotic cells. MPKs are responsible for phosphorylation and inactivation of the branched-chain alpha-ketoacid dehydrogenase and pyruvate dehydrogenase complexes (Harris, R.A. et al. (1995) Adv. Enzyme Regul. 34:147-162). Five MPKs have been identified. Four members correspond to pyruvate dehydrogenase kinase isozymes, regulating the activity of the pyruvate dehydrogenase complex, which is an important regulatory enzyme at the interface between glycolysis and the citric acid cycle. The fifth member corresponds to a branched-chain alpha-ketoacid dehydrogenase kinase, important in the regulation of the pathway for the disposal of branched-chain amino acids. (Harris, R.A. et al. (1997) Adv. Enzyme Regul.
37:271-293). Both starvation and the diabetic state are known to result in a great increase in the activity of the pyruvate dehydrogenase kinase in the liver, heart and muscle of the rat. This increase contributes in both disease states to the phosphorylation and inactivation of the pyruvate dehydrogenase complex and conservation of pyruvate and lactate for gluconeogenesis (Harris (1995) su ra).
KINASES WITH NON-PROTEIN SUBSTRATES
Liyid and Inositol kinases Lipid kinases phosphorylate hydroxyl residues on lipid head groups. A family of kinases involved in phosphorylation of phosphatidylinositol (PI) has been described, each member l0 phosphorylating a specific carbon on the inositol ring (Leevers, S.J. et al. (1999) Curr. Opin. Cell. Biol.
11:219-225). The phosphorylation of phosphatidylinositol is involved in activation of the protein kinase C signaling pathway. The inositol phospholipids (phosphoinositides) intracellular signaling pathway begins with binding of a signaling molecule to a G-protein linked receptor in the plasma membrane.
This leads to the phosphorylation of phosphatidylinositol (Pl) residues on the inner side of the plasma membrane by inositol kinases, thus converting PI residues to the biphosphate state (PIPS. PIPZ is then cleaved into inositol triphosphate (IP3) and diacylglycerol. These two products act as mediators for separate signaling pathways. Cellular responses that are mediated by these pathways are glycogen breakdown in the liver in response to vasopressin, smooth muscle contraction in response to acetylcholine, and thrombin-induced platelet aggregation.
PI 3-kinase (PI3K), which phosphorylates the D3 position of PI and its derivatives, has a ' central role in growth factor signal cascades involved in cell growth, differentiation, and metabolism.
PI3K is a heterodimer consisting of an adapter subunit and a catalytic subunit. The adapter subunit acts as a scaffolding protein, interacting with specific tyrosine-phosphorylated proteins, lipid moieties, and other cytosolic factors. When the adapter subunit binds tyrosine phosphorylated targets, such as the insulin responsive substrate (IRS)-l, the catalytic subunit is activated and converts PI (4,5) bisphosphate (PIPS to PI (3,4,5) P3 (PIPS). PIPS then activates a number of other proteins, including PKA, protein kinase B (PKB), protein kinase C (PKC), glycogen synthase kinase (GSK)-3, and p70 ribosomal s6 kinase. PI3K also interacts directly with the cytoskeletal organizing proteins, Rac, rho, and cdc42 (Shepherd, P.R. et al. (1998) Biochem. J. 333:471-490). Animal models for diabetes, such as obese and fat mice, have altered PI3K adapter subunit levels. Specific mutations in the adapter subunit have also been found in an insulin-resistant Danish population, suggesting a role for PI3K in type-2 diabetes (Shepard, supra).
An example of lipid kinase phosphorylation activity is the phosphorylation of l0 D-erythro-sphingosine to the sphingolipid metabolite, sphingosine-1-phosphate (SPP). SPP has emerged as a novel lipid second-messenger. with both extracellular and intracellular actions (Kohama, T. et al. (1998) J. Biol. Chem. 273:23722-23728). Extracellularly, SPP is a ligand for the G-protein coupled receptor EDG-1 (endothelial-derived, G-protein coupled receptor).
Intracellularly, SPP
regulates cell growth, survival, motility, and cytoskeletal changes. SPP
levels are regulated by sphingosine kinases that specifically phosphorylate D-erythro-sphingosine to SPP. The importance of sphingosine kinase in cell signaling is indicated by the fact that various stimuli, including platelet-derived growth factor (PDGF), nerve growth factor, and activation of protein kinase C, increase cellular levels of SPP by activation of sphingosine kinase, and the fact that competitive l0 inhibitors of the enzyme selectively inhibit cell proliferation induced by PDGF (Kohama et al., supra).
Purine Nucleotide Kinases The purine nucleotide kinases, adenylate kinase (ATP:AMP phosphotransferase, or AdK) and guanylate kinase (ATP:GMP phosphotransferase, or GuK) play a key role in nucleotide metabolism and are crucial to the synthesis and regulation of cellular levels of ATP and GTP, respectively. These two molecules are precursors in DNA and RNA synthesis in growing cells and provide the primary source.of biochemical energy in cells (ATP), and signal transduction pathways (GTP). Inhibition of various steps in the synthesis of these two molecules has been the basis of many antiproliferative drugs for cancer and antiviral therapy (Pillwein, K. et al. (1990) Cancer Res.
50:1576-.1579).
AdK is found in almost all cell types and is especially abundant in cells having high rates of ATP synthesis and utilization such as skeletal muscle. In these cells AdK is physically associated with mitochondria and myofibrils, the subcellulax structures that are involved in energy production and utilization, respectively. Recent studies have demonstrated a major function for AdK in transferring high energy phosphoryls from metabolic processes generating ATP to cellular components consuming ATP (Zeleznikar, R.J. et al. (1995) J. Biol. Chem. 270:7311-7319). Thus AdK
may have a pivotal role in maintaining energy production in cells, particularly those having a high rate of growth or metabolism such as cancer cells, and may provide a target for suppression of its activity to treat certain cancers. Alternatively, reduced AdK activity may be a source of various metabolic, muscle-energy disorders that can result in cardiac or respiratory failure and may be treatable by increasing AdK activity.
GuK, in addition to providing a key step in the synthesis of GTP for RNA and DNA synthesis, also fulfills an essential function in signal transduction pathways of cells through the regulation of GDP
and GTP. Specifically, GTP binding to membrane associated G proteins mediates the activation of cell receptors, subsequent intracellular activation of adenyl cyclase, and production of the second il messenger, cyclic AMP. GDP binding to G proteins inhibits these processes. GDP
and GTP levels also control the activity of certain oncogenic proteins such as p21~ known to be involved in control of cell proliferation and oncogenesis (Bos, J.L. (1989) Cancer Res. 49:4682-4689). High ratios of GTP:GDP caused by suppression of GuK cause activation of p21'~ and promote oncogenesis.
Increasing GuK activity to increase levels of GDP and reduce the GTP:GDP ratio may provide a therapeutic strategy to reverse oncogenesis.
GuK is an important enzyme in the phosphorylation and activation of certain antiviral drugs useful in the treatment of herpes virus infections. These drugs include the guanine homologs acyclovir and buciclovir (Miller, W.H. and R.L. Miller (1980) J. Biol. Chem. 255:7204-7207; Stenberg, K. et al.
(1986) J. Biol. Chem. 261:2134-2139). Increasing GuK activity in infected cells may provide a therapeutic strategy for augmenting the effectiveness of these drugs and possibly for reducing the necessary dosages of the drugs.
Pyrimidine Kinases The pyrimidine kinases are deoxycytidine kinase and thymidine kinase 1 and 2.
Deoxycytidine kinase is located in the nucleus, and thymidine kinase 1 and 2 are found in the cytosol (Johansson, M.
et al. (1997) Proc. Natl. Acad. Sci. USA 94:11941-11945). Phosphorylation of deoxyribonucleosides by pyrimidine kinases provides an alternative pathway for de novo synthesis of DNA precursors. The role of pyrimidine kinases, like purine kinases, in phosphorylation is critical to the activation of several chemotherapeutically important nucleoside analogues (Arner E.S. and S.
Eriksson (1995) Pharmacol.
Ther.67:155-186).
The discovery of new human kinases, and the polynucleotides encoding them, satisfies a need in the art by providing new compositions which are useful in the diagnosis, prevention, and treatment of cancer, immune disorders, disorders affecting growth and development, cardiovascular diseases, and lipid disorders, and in the assessment of the effects of exogenous compounds on the expression of nucleic acid and amino acid sequences of human kinases.
SUMMARY OF THE INVENTION
The invention features purified polypeptides, human kinases, referred to collectively as "PKIN" and individually as "PKIN-1," "PKIN-2," "PKIN-3," "PKIN-4," "PKIN-5,"
"PKIN-6,"
"PKIN-7," "PKIN-8," "PKIN-9," "PKIN-10," "PKIN-11," "PKIN-12°' "PKIN-13," "PKIN-14,>' "PKIN-15," "PKIN-16," "PKI1V-17," "PKIIV-18," "PKIN-19," and "PKIIV-20." In one aspect, the invention provides an isolated polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID
NO:1-20, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90%
identical to an amino acid sequence selected from the group consisting of SEQ ID N0:1-20, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ
ID NO:1-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:l-20. In one alternative, the invention provides an isolated polypeptide comprising the amino acid sequence of SEQ ID N0:1-20.
The invention further provides an isolated polynucleotide encoding a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID N0:1-20, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:l-20, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID N0:1-20. In one alternative, the polynucleotide encodes a polypeptide selected from the group consisting of SEQ
ID N0:1-20. In another alternative, the polynucleotide is selected from the group consisting of SEQ ID N0:21-40.
Additionally, the invention provides a recombinant polynucleotide comprising a promoter sequence operably linked to a polynucleotide encoding a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:l-20, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID N0:1-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID N0:1-20. In one alternative, the invention provides a cell transformed with the recombinant polynucleotide. In another alternative, the invention provides a transgenic organism comprising the recombinant polynucleotide.
The invention also provides a method for producing a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID N0:1-20, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID N0:1-20, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:l-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID N0:1-20. The method comprises a) culturing a cell under conditions suitable for expression of the polypeptide, wherein said cell is transformed with a recombinant polynucleotide comprising a promoter sequence operably finked to a polynucleotide encoding the polypeptide, and b) recovering the polypeptide so expressed.
Additionally, the invention provides an isolated antibody which specifically binds to a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, c) a biologically active fragment of a polypeptide having.an amino acid~sequence selected from the group consisting of SEQ ID NO:1-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ
l0 ID NO:1-20.
The invention further provides an isolated polynucleotide selected from the group consisting of a) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ
ID N0:21-40, b) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90% identical to a polynucleotide sequence selected from the group consisting of SEQ ID N0:21-40, c) a polynucleotide complementary to the polynucleotide of a), d) a polynucleotide complementary to the polynucleotide of b), and e) an RNA equivalent of a)-d). In one alternative, the polynucleotide comprises at least 60 contiguous nucleotides.
Additionally, the invention provides a method for detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide selected from the group consisting of a) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ
ID N0:21-40, b) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90% identical to a polynucleotide sequence selected from the group consisting of SEQ ID N0:21-40, c) a polynucleotide complementary to the polynucleotide of a), d) a polynucleotide complementary to the polynucleotide of b), and e) an RNA equivalent of a)-d). The method comprises a) hybridizing the sample with a probe comprising at least 20 contiguous nucleotides comprising a sequence complementary to said target polynucleotide in the sample, and which probe specifically hybridizes to said target polynucleotide, under conditions whereby a hybridization complex is formed between said probe and said target polynucleotide or fragments thereof, and b) detecting the presence or absence of said hybridization complex, and optionally, if present, the amount thereof. In one alternative, the probe comprises at least 60 contiguous nucleotides.
The invention further provides a method for detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide selected from the group consisting of a) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ ID

N0:21-40, b) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90%
identical to a polynucleotide sequence selected from the group consisting of SEQ ID N0:21-40, c) a polynucleotide complementary to the polynucleotide of a), d) a polynucleotide complementary to the polynucleotide of b), and e) an RNA equivalent of a)-d). The method comprises a) amplifying said target polynucleotide or fragment thereof using polymerase chain reaction amplification, and b) detecting the presence or absence of said amplified target polynucleotide or fragment thereof, and, optionally, if present, the amount thereof.
The invention fiufiher provides a composition comprising an effective amount of a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID N0:1-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, and a pharmaceutically acceptable excipient. In one embodiment, the composition comprises an amino acid sequence selected from the group consisting of SEQ ID NO:l-20. The invention additionally provides a method of treating a disease or condition associated with decreased expression of functional PKIN, comprising administering to a patient in need of such treatment the composition.
The invention also provides a method for screening a compound for effectiveness as an , agonist of a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID N0:1-20, b) a polypeptide comprising a naturally occurring amino acid sequence at Least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID
NO:l-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ~ NO:1-20. The method comprises a) exposing a sample comprising the polypeptide to a compound, and b) detecting agonist activity in the sample. In one alternative, the invention provides a composition comprising an agonist compound identified by the method and a pharmaceutically acceptable excipient. In another alternative, the invention provides a method of treating a disease or condition associated with decreased expression of functional PKIN, comprising administering to a patient in need of such treatment the composition.
Additionally, the invention provides a method for screening a compound for effectiveness as an antagonist of a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID N0:1-20, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:l-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID N0:1-20. The method comprises a) exposing a sample comprising the polypeptide to a compound, and b) detecting antagonist activity in the sample.
In one alternative, the invention provides a composition comprising an antagonist compound identified by the method and a pharmaceutically acceptable excipient. In another alternative, the invention provides a method of treating a disease or condition associated with overexpression of functional PKIN, comprising administering to a patient in need of such treatment the composition.
The invention further provides a method of screening for a compound that specifically binds to a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ll~ N0:1-20, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ iD NO:1-20, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID
NO:I-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ )D NO:l-20. The method comprises a) combining the polypeptide with at least one test compound under suitable conditions, and b) detecting binding of the polypeptide to the test compound, thereby identifying a compound that specifically binds to the polypeptide.
The invention further provides a method of screening for a compound that modulates the activity of a polypeptide selected from the group consisting of a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ m N0:1-20, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ~ NO:1-20, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID
NO:1-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ m N0:1-20. The method comprises a) combining the polypeptide with at least one test compound under conditions permissive for the activity of the polypeptide, b) assessing the activity of the polypeptide in the presence of the test compound, and c) comparing the activity of the polypeptide in the presence of the test compound with the activity of the polypeptide in the absence of the test compound, wherein a change in the activity of the polypeptide in the presence of the test compound is indicative of a compound that modulates the activity of the polypeptide.
The invention further provides a method for screening a compound for effectiveness in altering expression of a target polynucleotide, wherein said target polynucleotide comprises a polynucleotide sequence selected from the group consisting of SEQ ID N0:21-40, the method comprising a) exposing a sample comprising the target polynucleotide to a compound, and b) detecting altered expression of the target polynucleotide.
The invention further provides a method for assessing toxicity of a test compound, said method comprising a) treating a biological sample containing nucleic acids with the test compound; b) hybridizing the nucleic acids of the treated biological sample with a probe comprising at least 20 contiguous nucleotides of a polynucleotide selected from the group consisting of i) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ
ID N0:21-40, ii) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90% identical to a polynucleotide sequence selected from the group consisting of SEQ ~ N0:21-40, iii) a polynucleotide having a sequence complementary to i), iv) a polynucleotide complementary to the polynucleotide of ii), and v) an RNA equivalent of i)-iv). Hybridization occurs under conditions whereby a specific hybridization complex is formed between said probe and a target polynucleotide in the biological sample, said target polynucleotide selected from the group consisting of i) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ ID N0:21-40, ii) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90% identical to a polynucleotide sequence selected from the group consisting of SEQ ID NO:21-40, iii) a polynucleotide complementary to the polynucleotide of i), iv) a polynucleotide complementary to the polynucleotide of ii), and v) an RNA equivalent of i)-iv). Alternatively, the target polynucleotide comprises a fragment of a polynucleotide sequence selected from the group consisting of i)-v) above; c) quantifying the amount of hybridization complex; and d) comparing the amount of hybridization complex in the treated biological sample with the amount of hybridization complex in an untreated biological sample, wherein a difference in the amount of hybridization complex in the treated biological sample is indicative of toxicity of the test compound.
BRIEF DESCRIPTION OF THE TABLES
Table 1 summarizes the nomenclature for the full length polynucleotide and polypeptide sequences of the present invention.
Table 2 shows the GenBank identification number and annotation of the nearest GenBank homolog for polypeptides of the invention. The probability score for the match between each polypeptide and its GenBankhomolog is also shown.
Table 3 shows structural features of polypeptide sequences of the invention, including predicted motifs and domains, along with the methods, algorithms, and searchable databases used for analysis of the polypeptides.
Table 4 lists the cDNA and/or genomic DNA fragments which were used to assemble polynucleotide sequences of the invention, along with selected fragments of the polynucleotide sequences.
Table 5 shows the representative cDNA library for polynucleotides of the invention.
Table 6 provides an appendix which describes the tissues and vectors used for construction of i0 the cDNA libraries shown in Table 5.
Table 7 shows the tools, programs, and algorithms used to analyze the polynucleotides and polypeptides of the invention, along with applicable descriptions, references, and threshold parameters.
DESCRIPTION OF THE INVENTION
Before the present proteins, nucleotide sequences, and methods are described, it is understood that this invention is not limited to the particular machines, materials and methods described, as these may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present invention which will be limited only by the appended claims.
It must be noted that as used herein and in the appended claims, the singular forms "a," "an,"
and "the" include plural reference unless the context clearly dictates otherwise. Thus, for example, a reference to "a host cell" includes a plurality of such host cells, and a reference to "an antibody" is a reference to one or more antibodies and equivalents thereof known to those skilled in the art, and so forth.
Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of ordinary skill in the art to which this invention belongs.
Although any machines, materials, and methods similar or equivalent to those described herein can be used to practice or test the present invention, the preferred machines, materials and methods are now described. All publications mentioned herein are cited for the purpose of describing and disclosing the cell lines, protocols, reagents and vectors which are reported in the publications and which might be used in connection with the invention. Nothing herein is to be construed as an admission that the invention is not entitled to antedate such disclosure by virtue of prior invention.
i~

DEFINITIONS
"PKIN" refers to the amino acid sequences of substantially purified PKIN
obtained from any species, particularly a mammalian species, including bovine, ovine, porcine, marine, equine, and human, and from any source, whether natural, synthetic, semi-synthetic, or recombinant.
The term "agonist" refers to a molecule which intensifies or mimics the biological activity of ,_ PKIN. Agonists may include proteins, nucleic acids, carbohydrates, small molecules, or any other compound or composition which modulates the activity of PKIN either by directly interacting with PKIN or by acting on components of the biological pathway in which PKIN
participates.
An "allelic variant" is an alternative form of the gene encoding PI~IN.
Allelic variants may result from at least one mutation in the nucleic acid sequence and may result in altered mRNAs or in polypeptides whose structure or function may or may not be altered. A gene may have none, one, or many allelic variants of its naturally occurring form. Common mutational changes which give rise to allelic variants are generally ascribed to natural deletions, additions, or substitutions of nucleotides.
Each of these types of changes may occur alone, or in combination with the others, one or more times in a given sequence.
"Altered" nucleic acid sequences encoding PKIN include those sequences with deletions, insertions, or substitutions of different nucleotides, resulting in a polypeptide the same as PI~IN or a polypeptide with at least one functional characteristic of PKIN. Included within this definition are polymorphisms which may or may not be readily detectable using a particular oligonucleotide probe of the polynucleotide encoding PI~IN, and improper or unexpected hybridization to allelic variants, with a locus other than the normal chromosomal locus for the polynucleotide sequence encoding PKIN. The encoded protein may also be "altered," and may contain deletions, insertions, or substitutions of amino acid residues which produce a silent change and result in a functionally equivalent PKIN. Deliberate amino acid substitutions may be made on the basis of similarity in polarity, charge, solubility, hydrophobicity, hydrophilicity, andlor the amphipathic nature of the residues, as long as the biological or immunological activity of PKIN is retained. For example, negatively charged amino acids may include aspartic acid and glutamic acid, and positively charged amino acids may include lysine and arginine. Amino acids with uncharged polar side chains having similar hydrophilicity values may include: asparagine and glutamine; and serine and threonine. Amino acids with uncharged side chains having similar hydrophilicity values may include: leucine, isoleucine, and valine; glycine and alanine;
and phenylalanine and tyrosine.
The terms "amino acid" and "amino acid sequence" refer to an oligopeptide, peptide, polypeptide, or protein sequence, or a fragment of any of these, and to naturally occurring or synthetic molecules. Where "amino acid sequence" is recited to refer to a sequence of a naturally occurring protein molecule, "amino acid sequence" and like terms are not meant to limit the amino acid sequence to the complete native amino acid sequence associated with the recited protein molecule.
"Amplification" relates to the production of additional copies of a nucleic acid sequence.
Amplification is generally carried out using polymerase chain reaction (PCR) technologies well known in the art.
The term "antagonist" refers to a molecule which inhibits or attenuates the biological activity of PI~IN. Antagonists may include proteins such as antibodies, nucleic acids, carbohydrates, small molecules, or any other compound or composition which modulates the activity of PKIN either by directly interacting with PHIN or by acting on components of the biological pathway in which PKIN
participates.
The term "antibody" refers to intact immunoglobulin molecules as well as to fragments thereof, such as Fab, F(ab')2, and Fv fragments, which are capable of binding an epitopic determinant.
Antibodies that bind PI~IN polypeptides can be prepared using intact polypeptides or using fragments containing small peptides of interest as the immunizing antigen. The polypeptide or oligopeptide used to immunize an animal (e.g., a mouse, a rat, or a rabbit) can be derived from the translation of RNA, or synthesized chemically, and can be conjugated to a carrier protein if desired. Commonly used carriers that are chemically coupled to peptides include bovine serum albumin, thyroglobulin, and keyhole limpet hemocyanin (KLI~. The coupled peptide is then used to immunize the animal.
The term "antigenic determinant" refers to that region of a molecule (i.e., an epitope) that makes contact with a particular antibody. When a protein or a fragment of a protein is used to immunize a host animal, numerous regions of the protein may induce the production of antibodies which bind specifically to antigenic determinants (particular regions or three-dimensional structures on the protein). An antigenic determinant may compete with the intact antigen (i.e., the immunogen used to elicit the immune response) for binding to an antibody.
The term "antisense" refers to any composition capable of base-pairing with the "sense"
(coding) strand of a specific nucleic acid sequence. Antisense compositions may include DNA; RNA;
peptide nucleic acid (PNA); oligonucleotides having modified backbone linkages such as phosphorothioates, methylphosphonates, or benzylphosphonates; oligonucleotides having modified sugar groups such as 2'-methoxyethyl sugars or 2'-methoxyethoxy sugars; or oligonucleotides having modified bases such as 5-methyl cytosine, 2'-deoxyuracil, or 7-deaza-2'-deoxyguanosine. Antisense molecules may be produced by any method including chemical synthesis or transcription. Once introduced into a cell, the complementary antisense molecule base-pairs with a naturally occurring nucleic acid sequence produced by the cell to form duplexes which block either transcription or translation. The designation "negative" or "minus" can refer to the antisense strand, and the designation "positive" or "plus" can refer to the sense strand of a reference DNA molecule.
The term "biologically active" refers to a protein having structural, regulatory, or biochemical functions of a naturally occurring molecule. Likewise, "immunologically active" or "immunogenic"
refers to the capability of the natural, recombinant, or synthetic PKIN, or of any oligopeptide thereof, to induce a specific immune response in appropriate animals or cells and to bind with specific antibodies.
"Complementary" describes the relationship between two single-stranded nucleic acid sequences that anneal by base-pairing. For example, 5'-AGT-3' pairs with its complement, 3'-TCA-5'.
A "composition comprising a given polynucleotide sequence" and a "composition comprising a given amino acid sequence" refer broadly to any composition containing the given polynucleotide or amino acid sequence. The composition may comprise a dry formulation or an aqueous solution.
Compositions comprising polynucleotide sequences encoding PKIN or fragments of PI~IN may be employed as hybridization probes. The probes may be stored in freeze-dried form and may be associated with a stabilizing agent such as a carbohydrate. In hybridizations, the probe may be deployed in an aqueous solution containing salts (e.g., NaCI), detergents (e.g., sodium dodecyl sulfate;
SDS), and other components (e.g., Denhardt's solution, dry milk, salmon sperm DNA, etc.).
"Consensus sequence" refers to a nucleic acid sequence which has been subjected to repeated DNA sequence analysis to resolve uncalled bases, extended using the XL-PCR kit (Applied Biosystems, Foster City CA) in the 5' and/or the 3' direction, and resequenced, or which has been assembled from one or more overlapping cDNA, EST, or genomic DNA fragments using a computer program for fragment assembly, such as the GELVIEW fragment assembly system (GCG, Madison WI) or Phrap (University of Washington, Seattle WA). Some sequences have been both extended and assembled to produce the consensus sequence.
"Conservative amino acid substitutions" are those substitutions that are predicted to least interfere with the properties of the original protein, i.e., the structure and especially the function of the protein is conserved and not significantly changed by such substitutions. The table below shows amino acids which may be substituted for an original amino acid in a protein and which are regarded as conservative amino acid substitutions.
Original Residue Conservative Substitution Ala Gly, Ser Arg His, Lys Asn Asp, Gln, His Asp Asn, Glu Cys Ala, Ser Gln Asn, Glu, His Glu Asp, Gln, His Gly Ala His Asn, Arg, Gln, Glu lle Leu, Val Leu Ile, Val Lys Arg, Gln, Glu Met Leu, Ile Phe ~ His, Met, Leu, Trp, Tyr Ser ~ Cys, Thr Thr Ser, Val Trp Phe, Tyr Tyr His, Phe, Trp Val Ile, Leu, Thr Conservative amino acid substitutions generally maintain (a) the structure of the polypeptide backbone in the area of the substitution, for example, as a beta sheet or alpha helical conformation, (b) the charge or hydrophobicity of the molecule at the site of the substitution, and/or (c) the bulk of the side chain.
A "deletion" refers to a change in the amino acid or nucleotide sequence that results in the absence of one or more amino acid residues or nucleotides.
The term "derivative" refers to a chemically modified polynucleotide or polypeptide.
Chemical modifications of a polynucleotide can include, for example, replacement of hydrogen by an alkyl, acyl, hydroxyl, or amino group. A derivative polynucleotide encodes a polypeptide which retains at least one biological or immunological function of the natural molecule. A
derivative polypeptide is one modified by glycosylation, pegylation, or any similar process that retains at least one biological or immunological function of the polypeptide from which it was derived.
A "detectable label" refers to a reporter molecule or enzyme that is capable of generating a measurable signal and is covalently or noncovalently joined to a polynucleotide or polypeptide.
"Differential expression" refers to increased or upregulated; or decreased, downregulated, or absent gene or protein expression, determined by comparing at least two different samples. Such comparisons may be carried out between, for example, a treated and an untreated sample, or a diseased and a normal sample.
"Exon shuffling" refers to the recombination of different coding regions (exons). Since an exon may represent a structural or functional domain of the encoded protein, new proteins may be assembled through the novel reassortment of stable substructures, thus allowing acceleration of the evolution of new protein functions.
A "fragment" is a unique portion of PKIN or the polynucleotide encoding PKIN
which is identical in sequence to but shorter in length than the parent sequence. A
fragment may comprise up to the entire length of the defined sequence, minus one nucleotidelamino acid residue. For example, a fragment may comprise from 5 to 1000 contiguous nucleotides or amino acid residues. A fragment used as a probe, primer, antigen, therapeutic molecule, or for other purposes, may be at least 5, 10, 15, 16, 20, 25, 30, 40, 50, 60, 75, 100, 150, 250 or at least 500 contiguous nucleotides or amino acid residues in length. Fragments may be preferentially selected from certain regions of a molecule. For example, a polypeptide fragment may comprise a certain length of contiguous amino acids selected l0 from the first 250 or 500 amino acids (or first 25% or 50%) of a polypeptide as shown in a certain defined sequence. Clearly these lengths are exemplary, and any length that is supported by the specification, including the Sequence Listing, tables, and figures, may be encompassed by the present embodiments.
A fragment of SEQ ID N0:21-40 comprises a region of unique polynucleotide sequence that specifically identifies SEQ ID N0:21-40, for example, as distinct from any other sequence in the genome from which the fragment was obtained. A fragment of SEQ ID N0:21-40 is useful, for example, in hybridization and amplification technologies and in analogous methods that distinguish SEQ
ID N0:21-40 from related polynucleotide sequences. The precise length of a fragment of SEQ ID
N0:21-40 and the region of SEQ ID N0:21-40 to which the fragment corresponds are routinely determinable by one of ordinary skill in the art based on the intended purpose for the fragment.
A fragment of SEQ ID NO:l-20 is encoded by a fragment of SEQ ID N0:21-40. A
fragment of SEQ ID NO:1-20 comprises a region of unique amino acid sequence that specifically identifies SEQ ID NO:1-20. For example, a fragment of SEQ ID N0:1-20 is useful as an immunogenic peptide for the development of antibodies that specifically recognize SEQ ID NO:1-20.
The precise length of a fragment of SEQ ID N0:1-20 and the region of SEQ ID
NO:1-20 to which the fragment corresponds are routinely determinable by one of ordinary skill in the art based on the intended purpose for the fragment.
A "full length" polynucleotide sequence is one containing at least a translation initiation codon (e.g., methionine) followed by an open reading frame and a translation termination codon. A "full length" polynucleotide sequence encodes a "full length" polypeptide sequence.
"Homology" refers to sequence similarity or, interchangeably, sequence identity, between two or more polynucleotide sequences or two or more polypeptide sequences.
The terms "percent identity" and "% identity," as applied to polynucleotide sequences, refer to the percentage of residue matches between at least two polynucleotide sequences aligned using a standardized algorithm. Such an algorithm may insert, in a standardized and reproducible way, gaps in the sequences being compared in order to optimize alignment between two sequences, and therefore achieve a more meaningful comparison of the two sequences.
Percent identity between polynucleotide sequences may be determined using the default parameters of the CLUSTAL V algorithm as incorporated into the MEGALIGN
version 3.12e sequence alignment program. This program is part of the LASERGENE software package, a suite of molecular biological analysis programs (DNASTAR, Madison Wn. CLUSTAL V is described in Higgins, D.G. and P.M. Sharp (1989) CABIOS 5:151-153 and in Higgins, D.G. et al. (1992) CABIOS
l0 8:189-191. For pairwise alignments of polynucleotide sequences, the default parameters are set as follows: Ktuple=2, gap penalty=5, window=4, and "diagonals saved"=4. The "weighted" residue weight table is selected as the default. Percent identity is reported by CLUSTAL V as the "percent similarity" between aligned polynucleotide sequences.
Alternatively, a suite of commonly used and freely available sequence comparison algorithms is provided by the National Center for Biotechnology Information (NCB17 Basic Local Alignment Search Tool (BLAST) (Altschul, S.F. et al. (1990) J. Mol. Biol. 215:403-410), which is available from several sources, including the NCBI, Bethesda, MD, and on the Internet at http://www.ncbi.nim.nih.govBLAST/. The BLAST software suite includes various sequence analysis programs including "blastn," that is used to align a known polynucleotide sequence with other polynucleotide sequences from a variety of databases. Also available is a tool called "BLAST 2 Sequences" that is used for direct pairwise comparison of two nucleotide sequences. "BLAST 2 Sequences" can be accessed and used interactively at http://www.ncbi.nlm.nih.gov/gorf/bl2.html. The "BLAST 2 Sequences" tool can be used for both blastn and blastp (discussed below). BLAST
programs are commonly used with gap and other parameters set to default settings. For example, to compare two nucleotide sequences, one may use blastn with the "BLAST 2 Sequences" tool Version 2Ø12 (April-21-2000) set at default parameters. Such default parameters may be, for example:
Matrix: BLOSUM62 Reward for match: 1 Penalty for »zismatclz: -2 Opera Gap: S afzd Extefzsiorz Gap: 2 penalties Gap x dt-op-off: 50 Expect: 10 Word Size: 11 Filter: on Percent identity may be measured over the length of an entire defined sequence, for example, as defined by a particular SEQ ID number, or may be measured over a shorter length, for example, over the length of a fragment taken from a larger, defined sequence, for instance, a fragment of at least 20, at least 30, at least 40, at least 50, at least 70, at least I00, or at least 200 contiguous nucleotides. Such lengths are exemplary only, and it is understood that any fragment length supported by the sequences shown herein, in the tables, figures, or Sequence Listing, may be used to describe a length over which percentage identity may be measured.
Nucleic acid sequences that do not show a high degree of identity may nevertheless encode similar amino acid sequences due to the degeneracy of the genetic code. It is understood that changes in a nucleic acid sequence can be made using this degeneracy to produce multiple nucleic acid sequences that all encode substantially the same protein.
The phrases "percent identity' and "% identity," as applied to polypeptide sequences, refer to the percentage of residue matches between at least two polypeptide sequences aligned using a i5 standardized algorithm. Methods of polypeptide sequence alignment are well-known. Some alignment methods take into account conservative amino acid substitutions. Such conservative substitutions, explained in more detail above, generally preserve the charge and hydrophobicity at the site of substitution, thus preserving the structure (and therefore function) of the polypeptide.
Percent identity between polypeptide sequences may be determined using the default parameters of the CLUSTAL V algorithm as incorporated into the MEGALIGN
version 3.12e sequence alignment program (described and referenced above). For pairwise alignments of polypeptide sequences using CLUSTAL V, the default parameters are set as follows: Ktuple=1, gap penalty=3, window=5, and "diagonals saved"=5. The PAM250 matrix is selected as the default residue weight table. As with polynucleotide alignments, the percent identity is reported by CLUSTAL V as the "percent similarity" between aligned polypeptide sequence pairs.
Alternatively the NCBI BLAST software suite may be used. For example, fox a pairwise comparison of two polypeptide sequences, one may use the "BLAST 2 Sequences"
tool Version 2Ø12 (April-21-2000) with blastp set at default parameters. Such default parameters may be, for example:
Matrix: BLOSUM62 Open Gap: ll ahd Extercsiofa Gap: I penalties Gap x drop-off SO
Expect: 10 Word Size: 3 Filter: ort Percent identity may be measured over the length of an entire defined polypeptide sequence, for example, as defined by a particular SEQ ID number, or may be measured over a shorter length, for example, over the length of a fragment taken from a larger, defined polypeptide sequence, for instance, a fragment of at least 15, at least 20, at least 30, at least 40, at least 50, at least 70 or at least 150 contiguous residues. Such lengths are exemplary only, and it is understood that any fragment length supported by the sequences shown herein, in the tables, figures or Sequence Listing, may be used to describe a length over which percentage identity may be measured.
"Human artificial chromosomes" (HACs) are linear microchromosomes which may contain DNA sequences of about 6 kb to 10 Mb in size and which contain all of the elements required for chromosome replication, segregation and maintenance.
The term "humanized antibody" refers to an antibody molecule in which the amino acid sequence in the non-antigen binding regions has been altered so that the antibody more closely resembles a human antibody, and still retains its original binding ability.
"Hybridization" refers to the process by which a polynucleotide strand anneals with a complementary strand through base pairing under defined hybridization conditions. Specific hybridization is an indication that two nucleic acid sequences share a high degree of complementarity.
Specific hybridization complexes form under permissive annealing conditions and remain hybridized after the "washing" step(s). The washing steps) is particularly important in determining the stringency of the hybridization process, with more stringent conditions allowing less non-specific binding, i.e., binding between pairs of nucleic acid strands that are not perfectly matched. Permissive conditions for annealing of nucleic acid sequences are routinely determinable by one of ordinary skill in the art and may be consistent among hybridization experiments, whereas wash conditions may be varied among experiments to achieve the desired stringency, and therefore hybridization specificity.
Permissive annealing conditions occur, for example, at 68°C in the presence of about 6 x SSC, about 1% (w/v) SDS, and about 100 ~g/ml sheared, denatured salmon sperm DNA.
Generally, stringency of hybridization is expressed, in part, with reference to the temperature under which the wash step is carried out. Such wash temperatures are typically selected to be about 5°C to 20°C lower than the thermal melting point (T~ for the specific sequence at a defined ionic strength and pH. The Tm is the temperature (under defined ionic strength and pH) at which 50% of the target sequence hybridizes to a perfectly matched probe. An equation for calculating Tm and conditions for nucleic acid hybridization are well known and can be found in Sambrook, J. et al. (1989) Molecular Cloning: A Laborator<~ Manual, 2"d ed., vol. 1-3, Cold Spring Harbor Press, Plainview NY;
specifically see volume 2, chapter 9.
High stringency conditions for hybridization between polynucleotides of the present invention include wash conditions of 68°C in the presence of about 0.2 x SSC and about 0.1% SDS, for 1 hour.
Alternatively, temperatures of about 65°C, 60°C, 55°C, or 42°C may be used. SSC concentration may be varied from about 0.1 to 2 x SSC, with SDS being present at about 0.1 %.
Typically, blocking reagents are used to block non-specific hybridization. Such blocking reagents include, for instance, sheared and denatured salmon sperm DNA at about 100-200 ~g/ml. Organic solvent, such as formamide at a concentration of about 35-50% v/v, may also be used under particular circumstances, such as for RNA:DNA hybridizations. Useful variations on these wash conditions will be readily apparent to those of ordinary skill in the art. Hybridization, particularly under high stringency conditions, may be suggestive of evolutionary similarity between the nucleotides. Such similarity is strongly indicative of a similar role for the nucleotides and their encoded polypeptides.
The term "hybridization complex" refers to a complex formed between two nucleic acid sequences by virtue of the formation of hydrogen bonds between complementary bases. A
hybridization complex may be formed in solution (e.g., Cot or Rot analysis) or formed between one nucleic acid sequence present in solution and another nucleic acid sequence immobilized on a solid support (e.g., paper, membranes, filters, chips, pins or glass slides, or any other appropriate substrate to which'cells or their nucleic acids have been fixed).
The words "insertion" and "addition" refer to changes in an amino acid or nucleotide sequence resulting in the addition of one or more amino acid residues or nucleotides, respectively.
"Immune response" can refer to conditions associated with inflammation, trauma, immune disorders, or infectious or genetic disease, etc. These conditions can be characterized by expression of various factors, e.g., cytokines, chemokines, and other signaling molecules, which may affect cellular and systemic defense systems.
An "immunogenic fragment" is a polypeptide or oligopeptide fragment of PKIN
which is capable of eliciting an immune response when introduced into a living organism, for example, a mammal. The term "immunogenic fragment" also includes any polypeptide or oligopeptide fragment of PI~IN which is useful in any of the antibody production methods disclosed herein or known in the art.
The term "microarray" refers to an arrangement of a plurality of polynucleotides, polypeptides, or other chemical compounds on a substrate.
The terms "element" and "array element" refer to a polynucleotide, polypeptide, or other chemical compound having a unique and defined position on a microarray.

The term "modulate" refers to a change in the activity of PKIN. For example, modulation may cause an increase or a decrease in protein activity, binding characteristics, or any other biological, functional, or immunological properties of PKIN.
The phrases "nucleic acid" and "nucleic acid sequence" refer to a nucleotide, oligonucleotide, polynucleotide, or any fragment thereof. These phrases also refer to DNA or RNA of genomic or synthetic origin which may be single-stranded or double-stranded and may represent the sense or the antisense strand, to peptide nucleic acid (PNA), or to any DNA-like or RNA-like material.
"Operably linked" refers to the situation in which a first nucleic acid sequence is placed in a functional relationship with a second nucleic acid sequence. For instance, a promoter is operably linked to a coding sequence if the promoter affects the transcription or expression of the coding sequence. Operably linked DNA sequences may be in close proximity or contiguous and, where necessary to join two protein coding regions, in the same reading frame.
"Peptide nucleic acid" (PNA) refers to an antisense molecule or anti-gene agent which comprises an oligonucleotide of at least about 5 nucleotides in length linked to a peptide backbone of amino acid residues ending in lysine. The terminal lysine confers solubility to the composition. PNAs preferentially bind complementary single stranded DNA or RNA and stop transcript elongation, and may be pegylated to extend their lifespan in the cell.
"Post-translational modification" of an PI~IN may involve lipidation, glycosylation, phosphorylation, acetylation, racemization, proteolytic cleavage, and other modifications known in the art. These processes may occur synthetically or biochemically. Biochemical modifications will vary by cell type depending on the enzymatic milieu of PKIN.
"Probe" refers to nucleic acid sequences encoding PKIN, their complements, or fragments thereof, which are used to detect identical, allelic or related nucleic acid sequences. Probes are isolated oligonucleotides or polynucleotides attached to a detectable label or reporter molecule.
Typical labels include radioactive isotopes, ligands, chemiluminescent agents, and enzymes. "Primers"
are short nucleic acids, usually DNA oligonucleotides, which may be annealed to a target polynucleotide by complementary base-pairing. The primer may then be extended along the target DNA strand by a DNA polymerise enzyme. Primer pairs can be used for amplification (and identification) of a nucleic acid sequence, e.g., by the polymerise chain reaction (PCR).
Probes and primers as used in the present invention typically comprise at least 15 contiguous nucleotides of a known sequence. In order to enhance specificity, longer probes and primers may also be employed, such as probes and primers that comprise at least 20, 25, 30, 40, 50, 60, 70, 80, 90, 100, or at least 150 consecutive nucleotides of the disclosed nucleic acid sequences. Probes and primers may be considerably longer than these examples, and it is understood that any length supported by the specification, including the tables, figures, and Sequence Listing, may be used.
Methods for preparing and using probes and primers are described in the references, for example Sambrook, J. et al. (1989) Molecular Cloning: A Laboratory Manual, 2"d ed., vol. 1-3, Cold Spring Harbor Press, Plainview NY; Ausubel, F.M. et al. (1987) Current Protocols in Molecular Biolo , Greene Publ. Assoc. & Wiley-Intersciences, New York NY; Innis, M. et al. (1990) PCR
Protocols, A Guide to Methods and Applications, Academic Press, San Diego CA.
PCR primer pairs can be derived from a known sequence, for example, by using computer programs intended for that purpose such as Primer (Version 0.5, 1991, Whitehead Institute for Biomedical Research, Cambridge io MA).
Oligonucleotides for use as primers are selected using software known in the art for such purpose. For example, OLIGO 4.06 software is useful for the selection of PCR
primer pairs of up to 100 nucleotides each, and for the analysis of oligonucleotides and larger polynucleotides of up to 5,000 nucleotides from an input polynucleotide sequence of up to 32 kilobases.
Similar primer selection IS programs have incorporated additional features for expanded capabilities.
For example, the PrimOU
primer selection program (available to the public from the Genome Center at University of Texas South West Medical Center, Dallas TX) is capable of choosing specific primers from megabase sequences and is thus useful for designing primers on a genome-wide scope. The Primer3 primer selection program (available to the public from the Whitehead Institute/MIT
Center for Genome 20 Research, Cambridge MA) allows the user to input a "mispriming library," in which sequences to avoid as primer binding sites are user-specified. Primer3 is useful, in particular, for the selection of oligonucleotides for microarrays. (The source code for the latter two primer selection programs may also be obtained from their respective som.~ces and modified to meet the user's specific needs.) The PrimeGen program (available to the public from the UK Human Genome Mapping Project Resource 25 Centre, Cambridge UK) designs primers based on~multiple sequence alignments, thereby allowing selection of primers that hybridize to either the most conserved or least conserved regions of aligned nucleic acid sequences. Hence, this program is useful for identification of both unique and conserved oligonucleotides and polynucleotide fragments. The oligonucleotides and polynucleotide fragments identified by any of the above selection methods are useful in hybridization technologies, for example, 30 as PCR or sequencing primers, microarray elements, or specific probes to identify fully or partially complementary polynucleotides in a sample of nucleic acids. Methods of oligonucleotide selection are not limited to those described above.
A "recombinant nucleic acid" is a sequence that is not naturally occurring or has a sequence that is made by an artificial combination of two or more otherwise separated segments of sequence.
This artificial combination is often accomplished by chemical synthesis or, more commonly, by the artificial manipulation of isolated segments of nucleic acids, e.g., by genetic engineering techniques such as those described in Sambrook, sue. The term recombinant includes nucleic acids that have been altered solely by addition, substitution, or deletion of a portion of the nucleic acid. Frequently, a recombinant nucleic acid may include a nucleic acid sequence operably linked to a promoter sequence.
Such a recombinant nucleic acid may be part of a vector that is used, for example, to transform a cell.
Alternatively, such recombinant nucleic acids may be part of a viral vector, e.g., based on a vaccinia virus, that could be use to vaccinate a mammal wherein the recombinant nucleic acid is expressed, inducing a protective immunological response in the mammal.
A "regulatory element" refers to a nucleic acid sequence usually derived from untranslated regions of a gene and includes enhancers, promoters, introns, and 5' and 3' untranslated regions (UTRs). Regulatory elements interact with host or viral proteins which control transcription, translation, or RNA stability.
"Reporter molecules" are chemical or biochemical moieties used for labeling a nucleic acid, amino acid, or antibody. Reporter molecules include radionuclides; enzymes;
fluorescent, chemiluminescent, or chromogenic agents; substrates; cofactors; inhibitors;
magnetic particles; and other moieties known in the art.
An "RNA equivalent," in reference to a DNA sequence, is composed of the same lineax sequence of nucleotides as the reference DNA sequence with the exception that all occurrences of the nitrogenous base thymine are replaced with uracil, and the sugar backbone is composed of ribose instead of deoxyribose.
The term "sample" is used in its broadest sense. A sample suspected of containing PKIN, nucleic acids encoding PKIN, or fragments thereof may comprise a bodily fluid;
an extract from a cell, chromosome, organelle, or membrane isolated from a cell; a cell; genomic DNA, RNA, or cDNA, in solution or bound to a substrate; a tissue; a tissue print; etc.
The terms "specific binding" and "specifically binding" refer to that interaction between a protein or peptide and an agonist, an antibody, an antagonist, a small molecule, or any natural or synthetic binding composition. The interaction is dependent upon the presence of a particular structure of the protein, e.g., the antigenic determinant or epitope, recognized by the binding molecule. For example, if an antibody is specific for epitope "A," the presence of a polypeptide comprising the epitope A, or the presence of free unlabeled A, in a reaction containing free labeled A a.nd the antibody will reduce the amount of labeled A that binds to the antibody.

The term "substantially purified" refers to nucleic acid or amino acid sequences that are removed from their natural environment and are isolated or separated, and are at least 60% free, preferably at least 75% free, and most preferably at least 90% free from other components with which they are naturally associated.
A "substitution" refers to the replacement of one or more amino acid residues or nucleotides by different amino acid residues or nucleotides, respectively.
"Substrate" refers to any suitable rigid or semi-rigid support including membranes, filters, chips, slides, wafers, fibers,' magnetic or nonmagnetic beads, gels, tubing, plates, polymers, microparticles and capillaries. The substrate can have a variety of surface forms, such as wells, trenches, pins, channels and poxes, to which polynucleotides or polypeptides are bound.
A "transcript image" refers to the collective pattern of gene expression by a particular cell type or tissue under given conditions at a given time.
"Transformation" describes a process by which exogenous DNA is introduced into a recipient cell. Transformation may occur under natural or artificial conditions according to various methods IS well known in the art, and may rely on any known method for the insertion of foreign nucleic acid sequences into a prokaryotic or eukaryotic host cell. The method for transformation is selected based on the type of host cell being transformed and may include, but is not limited to, bacteriophage or viral infection, electroporation, heat shock, lipofection, and particle bombardment.
The term "transformed cells" includes stably transformed cells in which the inserted DNA is capable of replication either as an autonomously replicating plasmid or as part of the host chromosome, as well as transiently transformed cells which express the inserted DNA or RNA for limited periods of time.
A "transgenic organism," as used herein, is any organism, including but not limited to animals and plants, in which one or more of the cells of the organism contains heterologous nucleic acid introduced by way of human intervention, such as by transgenic techniques well known in the art. The nucleic acid is introduced into the cell, directly or indirectly by introduction into a precursor of the cell, by way of deliberate genetic manipulation, such as by microinjection or by infection with a recombinant virus. The term genetic manipulation does not include classical cross-breeding, or in vitro fertilization, but rather is directed to the introduction of a recombinant DNA
molecule. The transgenic organisms contemplated in accordance with the present invention include bacteria, cyanobacteria, fungi, plants and animals. The isolated DNA of the present invention can be introduced into the host by methods known in the art, for example infection, transfection, transformation or transconjugation.
Techniques for transfernng the DNA of the present invention into such organisms are widely known and provided in references such as Sambrook et al. (I989), supra.

A "variant" of a particular nucleic acid sequence is defined as a nucleic acid sequence having at least 40% sequence identity to the particular nucleic acid sequence over a certain length of one of the nucleic acid sequences using blastn with the "BLAST 2 Sequences" tool Version 2Ø9 (May-07-1999) set at default parameters. Such a pair of nucleic acids may show, for example, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% or greater sequence identity over a certain defined length. A variant may be described as, for example, an "allelic" (as defined above), "splice," "species," or "polymorphic" variant. A
splice variant may have significant identity to a reference molecule, but will generally have a greater or lesser number of polynucleotides due to alternate splicing of exons during mRNA processing. The corresponding polypeptide may possess additional functional domains or lack domains that are present in the reference molecule. Species variants are polynucleotide sequences that vary from one species to another. The resulting polypeptides will generally have significant amino acid identity relative to each other. A polymorphic variant is a variation in the polynucleotide sequence of a particular gene between individuals of a given species. Polymorphic variants also may encompass "single nucleotide polymorphisms" (SNPs) in which the polynucleotide sequence varies by one nucleotide base. The presence of~SNPs may be indicative of, for example, a certain population, a disease state, or a propensity for a disease state.
A "variant" of a particular polypeptide sequence is defined as a polypeptide sequence having at least 40% sequence identity to the particular polypeptide sequence over a certain length of one of the polypeptide sequences using blastp with the "BLAST 2 Sequences" tool Version 2Ø9 (May-07-1999) set at default parameters. Such a pair of polypeptides may show, for example, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%
or greater sequence identity over a certain defined length of one of the polypeptides.
THE INVENTION
The invention is based on the discovery of new human human kinases (PKIN), the polynucleotides encoding PKIN, and the use of these compositions for the diagnosis, treatment, or prevention of cancer, immune disorders, disorders affecting growth and development, cardiovascular diseases, and lipid disorders.
Table 1 summarizes the nomenclature for the full length polynucleotide and polypeptide sequences of the invention. Each polynucleotide and its corresponding polypeptide are correlated to a single Incyte project identification number (Incyte Project ID). Each polypeptide sequence is denoted by both a polypeptide sequence identification number (Polypeptide SEQ ID NO:) and an Incyte polypeptide sequence number (Incyte Polypeptide 1D) as shown. Each polynucleotide sequence is denoted by both a polynucleotide sequence identification number (Polynucleotide SEQ ID NO:) and an Incyte polynucleotide consensus sequence number (Incyte Polynucleotide 1D) as shown.
Table 2 shows sequences with homology to the polypeptides of the invention as identified by BLAST analysis against the GenBank protein (genpept) database. Columns 1 and 2 show the polypeptide sequence identification number (Polypeptide SEQ ID NO:) and the corresponding Incyte polypeptide sequence number (Incyte Polypeptide 1D) for polypeptides of the invention. Column 3 shows the GenBank identification number (Genbank ID NO:) of the nearest GenBank homolog.
Column 4 shows the probability score for the match between each polypeptide and its GenBank homolo8. Column 5 shows the annotation of the GenB ank homolog along with relevant citations where applicable, all of which are expressly incorporated by reference herein.
Table 3 shows various structural features of the polypeptides of the invention. Columns 1 and 2 show the polypeptide sequence identification number (SEQ ID NO:) and the corresponding Incyte polypeptide sequence number (Incyte Polypeptide ID) for each polypeptide of the invention. Column 3 shows the number of amino acid residues in each polypeptide. Column 4 shows potential phosphorylation sites, and column 5 shows potential glycosylation sites, as determined by the MOTIFS
program of the GCG sequence analysis software package (Genetics Computer Group, Madison WI).
Column 6 shows amino acid residues comprising signature sequences, domains, and motifs. Column 7 shows analytical methods for protein structure/function analysis and in some cases, searchable databases to which the analytical methods were applied.
Together, Tables 2 and 3 summarize the properties of polypeptides of the invention, and these properties establish that the claimed polypeptides are human kinases. For example, SEQ ID N0:2 is 97% identical to mouse tousled-like kinase (GenBank ID 82853031) as determined by the Basic Local Alignment Search Tool (BLAST). (See Table 2.) The BLAST probability score is 0.0, which indicates the probability of obtaining the observed polypeptide sequence alignment by chance. SEQ
ID N0:2 also contains an eukaryotic protein kinase domain as determined by searching for statistically significant matches in the hidden Markov model (HMM)-based PFAM database of conserved protein family domains. (See Table 3.) Data from BLIMPS, MOTIFS, and PROFILESCAN
analyses provide further corroborative evidence that SEQ ID N0:2 is a tousled-like kinase. In an alternative example, SEQ ID NO:10 is 63 % identical to human serine/threonine protein kinase (GenB ank ID
836615) as determined by the Basic Local Alignment Search Tool (BLAST). (See Table 2.) The BLAST probability score is 7.7e-122, which indicates the probability of obtaining the observed polypeptide sequence alignment by chance. SEQ ID N0:10 also contains an eukaryotic protein kinase domain as determined by searching for statistically significant matches in the hidden Markov model (HMM)-based PFAM database of conserved protein domains. (See Table 3.) Data from BLIMPS, MOTIFS, and PROFILESCAN analyses provide further corroborative evidence that SEQ ID NO:10 is a serine/threonine kinase. Note that "serine/theronine kinase" is a specific class of kinases. In an alternative example, SEQ ID N0:16 is 53 % identical to human receptor protein-tyrosine kinase (GenBank ID 8551608) as determined by the Basic Local Alignment Search Tool (BLAST). (See Table 2.) The BLAST probability score is 4.1e-290, which indicates the probability of obtaining the observed polypeptide sequence alignment by chance. SEQ ID N0:16 also contains an eukaryotic protein kinase domain as determined by searching for statistically significant matches in the hidden Markov model (HMM)-based PFAM database of conserved protein family domains.
.(See Table 3.) Data from BLIMPS and PROFILESCAN analyses provide further corroborative evidence that SEQ
ID N0:16 is a receptor tyrosine kinase. In an alternative example, SEQ ID
N0:19 is 93% identical to rat Calcium/calmodulin-dependent protein kinase isoform IV (GenBank ID
81836161) as determined by the Basic Local Alignment Search Tool (BLAST). (See Table 2.) The BLAST
probability score is 6.Oe-257, which indicates the probability of obtaining the observed polypeptide sequence alignment by chance. SEQ ID N0:19 also contains an eukaryotic protein kinase domain as determined by searching for statistically significant matches in the hidden Markov model (HMM)-based PFAM
database of conserved protein domains. (See Table 3.) Data from BLIMPS, MOTIFS, and PROFILESCAN analyses provide further corroborative evidence that SEQ ID N0:19 is a protein kinase. SEQ ID NO:1, SEQ ID N0:3-9, SEQ ID NO:11-15, SEQ ID N0:17-18, and SEQ
ID N0:20 were analyzed and annotated in a similar manner. The algorithms and parameters for the analysis of SEQ ID N0:1-20 are described in Table 7.
As shown in Table 4, the full length polynucleotide sequences of the present invention were assembled using cDNA sequences or coding (exon) sequences derived from genomic DNA, or any combination of these two types of sequences. Columns 1 and 2 list the polynucleotide sequence identification number (Polynucleotide SEQ ID NO:) and the corresponding Incyte polynucleotide consensus sequence number (Incyte Polynucleotide ID) for each polynucleotide of the invention.
Column 3 shows the len8th of each polynucleotide sequence in basepairs. Column 4 lists fragments of the polynucleotide sequences which are useful, for example, in hybridization or amplification technologies that identify SEQ ID N0:21-40 or that distinguish between SEQ ID
N0:21-40 and related polynucleotide sequences. Column 5 shows identification numbers corresponding to cDNA

sequences, coding sequences (exons) predicted from genomic DNA, and/or sequence assemblages comprised of both cDNA and genomic DNA. These sequences were used to assemble the full length polynucleotide sequences of the invention. Columns 6 and 7 of Table 4 show the nucleotide start (5') and stop (3') positions of the cDNA and/or genomic sequences in column 5 relative to their respective full length sequences.
The identification numbers in Column 5 of Table 4 may refer specifically, for example, to Incyte cDNAs along with their corresponding cDNA libraries. For example, 2564295H1 is the identification number of an Incyte cDNA sequence, and ADRETUT01 is the cDNA
library from which it is derived. Incyte cDNAs for which cDNA libraries are not indicated were derived from pooled cDNA libraries (e.g., 71191190V1). Alternatively, the identification numbers in column 5 may refer to GenBank cDNAs or ESTs (e.g., 81164223) which contributed to the assembly of the full length polynucleotide sequences. In addition, the identification numbers in column 5 may identify sequences derived from the ENSEMBL (The Sanger Centre, Cambridge, UI~) database (i.e., those sequences including the designation "ENST"). Alternatively, the identification numbers in column 5 may be derived from the NCBI RefSeq Nucleotide Sequence Records Database (i.
e., those sequences including the designation "NM" or "NT") or the NCBI RefSeq Protein Sequence Records (i.e., those sequences including the designation "NP"). Alternatively, the identification numbers in column 5 may refer to assemblages of both cDNA and Genscan-predicted exons brought together by an "exon stitching" algorithm. For example, FL XXXXXX NI NZ_YI'YYY N3 N.~
represents a "stitched" sequence in which X.~XXXX is the identification number of the cluster of sequences to which the algorithm was applied, and YYYYY is the number of the prediction generated by the algorithm, and N1,2,3...~ if present, represent specific exons that may have been manually edited during analysis (See Example V). Alternatively, the identification numbers in column 5 may refer to assemblages of exons brought together by an "exon-stretching" algorithm. For example, FT.~~X gAAA~9A_gBBBBB_1 Nis the identification number of a "stretched"
sequence, with XXXXXX being the Incyte project identification number, gAAAAA being the GenBank identification number of the human genomic sequence to which the "exon-stretching" algorithm was applied, gBBBBB being the GenBank identification number or NCBI RefSeq identification number of the nearest GenB ank protein homolog, and N referring to specific exons (See Example V). In instances where a RefSeq sequence was used as a protein homolog for the "exon-stretching" algorithm, a RefSeq identifier (denoted by "NM,"' "NP," or "NT") may be used in place of the GenBank identifier (i.e., gBBBBB).
Alternatively, a prefix identifies component sequences that were hand-edited, predicted from genomic DNA sequences, or derived from a combination of sequence analysis methods. The following Table lists examples of component sequence prefixes and corresponding sequence analysis methods associated with the prefixes (see Example IV and Example V).
Prefix Type of analysis andlor examples of programs GNN, GFG,Exon prediction from genomic sequences using, for example, ENST GENSCAN (Stanford University, CA, USA) or FGENES

(Computer Genomics Group, The Sanger Centre, Cambridge, UK) GBI Hand-edited analysis of genomic sequences.

FL Stitched or stretched genomic sequences (see Example V).

INCY Full length transcript and exon prediction from mapping of EST

sequences to the genome. Genomic location and EST composition data are combined to predict the exons and resulting transcript.

In some cases, Incyte cDNA coverage redundant with the sequence covetage shown in column 5 was obtained to confirm the final consensus polynucleotide sequence, but the relevant Incyte cDNA identification numbers are not shown.
Table 5 shows the representative cDNA libraries for those full length polynucleotide sequences which were assembled using Incyte cDNA sequences. The representative cDNA library is the Incyte cDNA library which is most frequently represented by the Incyte cDNA sequences which were used to assemble and confirm the above polynucleotide sequences.
The tissues and vectors which were used to construct the cDNA libraries shown in Table 5 are described in Table 6.
The invention also encompasses PKIN variants. A preferred PKIN variant is one which has at least about 80%, or alternatively at least about 90%, or even at least about 95% amino acid sequence identity to the PKIN amino acid sequence, and which contains at least one functional or structural characteristic of PKIN.
The invention also encompasses polynucleotides which encode PKIN. In a particular embodiment, the invention encompasses a polynucleotide sequence comprising a sequence selected from the group consisting of SEQ ID NO:21-40, which encodes PKIN. The polynucleotide sequences of SEQ ID N0:21-40, as presented in the Sequence Listing, embrace the equivalent RNA sequences, wherein occurrences of the nitrogenous base thymine are replaced with uracil, and the sugar backbone is composed of ribose instead of deoxyribose.
The invention also encompasses a variant of a polynucleotide sequence encoding PKIN. In particular, such a variant polynucleotide sequence will have at least about 70%, or alternatively at least about 85%, or even at least about 95% polynucleotide sequence identity to the polynucleotide sequence encoding PKIN. A particular aspect of the invention encompasses a variant of a polynucleotide sequence comprising a sequence selected from the group consisting of SEQ ID N0:21-40 which has at least about 70%, or alternatively at least about 85%, or even at least about 95%
polynucleotide sequence identity to a nucleic acid sequence selected from the group consisting of SEQ
ID N0:21-40. Any one of the polynucleotide variants described above can encode an amino acid sequence which contains at least one functional or structural characteristic of PKIN.
It will be appreciated by those skilled in the art that as a result of the degeneracy of the genetic code, a multitude of polynucleotide sequences encoding PKIN, some bearing minimal similarity to the polynucleotide sequences of any known and naturally occurring gene, may be produced. Thus, the invention contemplates each and every possible variation of polynucleotide sequence that could be made by selecting combinations based on possible codon choices. These combinations are made in accordance with the standard triplet genetic code as applied to the polynucleotide sequence of naturally occurring PK1N, and all such variations are to be considered as being specifically disclosed.
Although nucleotide sequences which encode PKIN and its variants are generally capable of hybridizing to the nucleotide sequence of the naturally occurring PKIN under appropriately selected conditions of stringency, it may be advantageous to produce nucleotide sequences encoding PKIN or its derivatives possessing a substantially different codon usage, e.g., inclusion of non-naturally occurring codons. Codons may be selected to increase the rate at which expression of the peptide occurs in a particular prokaryotic or eukaryotic host in accordance with the frequency with which particular codons are utilized by the host. Other reasons for substantially altering the nucleotide sequence encoding PKIN and its derivatives without altering the encoded amino acid sequences include the production of RNA transcripts having more desirable properties, such as a greater half life, than transcripts produced from the naturally occurring sequence.
The invention also encompasses production of DNA sequences which encode PK1N
and PKIN derivatives, or fragments thereof, entirely by synthetic chemistry. After production, the synthetic sequence may be inserted into any of the many available expression vectors and cell systems using reagents well known in the art. Moreover, synthetic chemistry may be used to introduce mutations into a sequence encoding PKIN or any fragment thereof.
Also encompassed by the invention are polynucleotide sequences that are capable of hybridizing to the claimed polynucleotide sequences, and, in particular, to those shown in SEQ ID
N0:21-40 and fragments thereof under various conditions of stringency. (See, e.g., Wahl, G.M. and S.L. Berger (1987) Methods Enzymol. 152:399-407; Kimmel, A.R. (1987) Methods Enzymol. 152:507-511.) Hybridization conditions, including annealing and wash conditions, are described in "Definitions."
Methods for DNA sequencing are well known in the art and may be used to practice any of the embodiments of the invention. The methods may employ such enzymes as the Klenow fragment of DNA polymerase I, SEQUENASE (US Biochemical, Cleveland OH), Taq polymerase (Applied Biosystems), thermostable T7 polymerase (Amersham Pharmacia Biotech, Piscataway NJ), or combinations of polymerases and proofreading exonucleases such as those found in the ELONGASE
amplification system (Life Technologies, Gaithersburg MD). Preferably, sequence preparation is automated with machines such as the MICROLAB 2200 liquid transfer system (Hamilton, Reno NV), PTC200 thermal cycler (MJ Research, Watertown MA) and ABI CATALYST 800 thermal cycler l0 (Applied Biosystems). Sequencing is then carried out using either the ABI
373 or 377 DNA
sequencing system (Applied Biosystems), the MEGABACE 1000 DNA sequencing system (Molecular Dynamics, Sunnyvale CA), or other systems known in the art. The resulting sequences are analyzed using a variety of algorithms which are well known in the art.
(See, e.g., Ausubel, F.M.
(1997) Short Protocols in Molecular Biolo~y, John Wiley & Sons, New York NY, unit 7.7; Meyers, R.A. (1995) Molecular Biology and Biotechnology, Wiley VCH, New York NY, pp.
856-853.) The nucleic acid sequences encoding PKIN may be extended utilizing a partial nucleotide sequence and employing various PCR-based methods known in the art to detect upstream sequences, such as promoters and regulatory elements. For example, one method which may be employed, restriction-site PCR, uses universal and nested primers to amplify unknown sequence from genomic DNA within a cloning vector. (See, e.g., Sarkar, G. .(1993) PCR Methods Applic. 2:318-322.) Another method, inverse PCR, uses primers that extend in divergent directions to amplify unknown sequence from a circularized template. The template is derived from restriction fragments comprising a known genomic locus and surrounding sequences. (See, e.g., Triglia, T. et al. (1988) Nucleic Acids Res. 16:8186.) A third method, capture PCR, involves PCR amplification of DNA
fragments adjacent to known sequences in human and yeast artificial chromosome DNA. (See, e.g., Lagerstrom, M. et al. (1991) PCR Methods Applic. 1:111-119.) Iu this method, multiple restriction enzyme digestions and ligations may be used to insert an engineered double-stranded sequence into a region of unknown sequence before performing PCR. Other methods which may be used to retrieve unknown sequences are known in the art. (See, e.g., Parker, J.D. et al. (1991) Nucleic Acids Res. 19:3055-3060).
Additionally, one may use PCR, nested primers, and PROMOTERFINDER libraries (Clontech, Palo Alto CA) to walk genomic DNA. This procedure avoids the need to screen libraries and is useful in finding intron/exon junctions. For all PCR-based methods, primers may be designed using commercially available software, such as OLIGO 4.06 primer analysis software (National Biosciences, Plymouth MN) or another appropriate program, to be about 22 to 30 nucleotides in length, to have a GC content of about 50% or more, and to anneal to the template at temperatures of about 68°C to 72°C.
When screening for full length cDNAs, it is preferable to use libraries that have been size-selected to include larger cDNAs. In addition, random-primed libraries, which often include sequences containing the 5' regions of genes, are preferable for situations in which an oligo d(T) library does not yield a full-length cDNA. Genomic libraries may be useful for extension of sequence into 5' non-transcribed regulatory regions.
Capillary electrophoresis systems which are commercially available may be used to analyze the size or confirm the nucleotide sequence of sequencing or PCR products. In particular, capillary sequencing may employ flowable polymers for electrophoretic separation, four different nucleotide specific, laser-stimulated fluorescent dyes, and a charge coupled device camera for detection of the emitted wavelengths. Output/light intensity may be converted to electrical signal using appropriate software (e.g., GENOTYPER and SEQUENCE NAVIGATOR, Applied Biosystems), and the entire process from loading of samples to computer analysis and electronic data display may be computer controlled. Capillary electrophoresis is especially preferable for sequencing small DNA fragments which may be present in limited amounts in a particular sample.
In another embodiment of the invention, polynucleotide sequences or fragments thereof which encode PKIN may be cloned in recombinant DNA molecules that direct expression of PKIN, or fragments or functional equivalents thereof, in appropriate host cells. Due to the inherent degeneracy of the genetic code, other DNA sequences which encode substantially the same or a functionally equivalent amino acid sequence may be produced and used to express PKIN.
The nucleotide sequences of the present invention can be engineered using methods generally known in the art in order to alter PI~IN-encoding sequences for a variety of purposes including, but not limited to, modification of the cloning, processing, and/or expression of the gene product. DNA
shuffling by random fragmentation and PCR reassembly of gene fragments and synthetic oligonucleotides may be used to engineer the nucleotide sequences. For example, oligonucleotide-mediated site-directed mutagenesis may be used to introduce mutations that create new restriction sites, alter glycosylation patterns, change codon preference, produce splice variants, and so forth.
The nucleotides of the present invention may be subjected to DNA shuffling techniques such as MOLECULARBREEDING (Maxygen Inc., Santa Clara CA; described in U.S. Patent Number 5,837,458; Chang, C.-C. et al. (1999) Nat. Biotechnol. 17:793-797; Christians, F.C. et al. (1999) Nat.
Biotechnol. 17:259-264; and Crameri, A. et al. (1996) Nat. Biotechnol. 14:315-319) to alter or improve the biological properties of PKIN, such as its biological or enzymatic activity or its ability to bind to other molecules or compounds. DNA shuffling is a process by which a library of gene variants is produced using PCR-mediated recombination of gene fragments. The library is then subjected to selection or screening procedures that identify those gene variants with the desired properties. These preferred variants may then be pooled and further subjected to recursive rounds of DNA shuffling and selection/screening. Thus, genetic diversity is created through "artificial"
breeding and rapid molecular evolution. For example, fragments of a single gene containing random point mutations may be recombined, screened, and then reshuffled until the desired properties are optimized. Alternatively, fragments of a given gene may be recombined with fragments of homologous genes in the same gene family, either from the same or different species, thereby maximizing the genetic diversity of multiple naturally occurring genes in a directed and controllable manner.
In another embodiment, sequences encoding PI~IN may be synthesized, in whole or in part, using chemical methods well known in the art. (See, e.g., Caruthers, M.H. et al. (1980) Nucleic Acids ' Symp. Ser. 7:215-223; and Horn, T. et al. (1980) Nucleic Acids Symp. Ser.
7:225-232.) Alternatively, PKIN itself or a fragment thereof may be synthesized using chemical methods.
For example, peptide synthesis can be performed using various solution-phase or solid-phase techniques. (See, e.g., Creighton, T. (1984) Proteins, Structures and Molecular Properties, WH
Freeman, New York NY, pp.
55-60; and Roberge, J.Y. et al. (1995) Science 269:202-204.) Automated synthesis may be achieved using the ABI 431A peptide synthesizer (Applied Biosystems). Additionally, the amino acid sequence of PKIN, or any part thereof, may be altered during direct synthesis and/or combined with sequences from other proteins, or any part thereof, to produce a variant polypeptide or a polypeptide having a sequence of a naturally occurring polypeptide.
The peptide may be substantially purified by preparative high performance liquid chromatography. (See, e.g., Chiez, R.M. and F.Z. Regnier (1990) Methods Enzymol. 182:392-421.) The composition of the synthetic peptides may be confirmed by amino acid analysis or by sequencing.
(See, e.g., Creighton, supra, pp. 28-53.) In order to express a biologically active PKIN, the nucleotide sequences encoding PI~IN or derivatives thereof may be inserted into an appropriate expression vector, i.e., a vector which contains the necessary elements for transcriptional and translational control of the inserted coding sequence in a suitable host. These elements include regulatory sequences, such as enhancers, constitutive and inducible promoters, and 5' and 3' untranslated regions in the vector and in polynucleotide sequences encoding PI~IN. Such elements may vary in their strength and specificity.
Specific initiation signals may also be used to achieve more efficient translation of sequences encoding PKIN. Such signals include the ATG initiation codon and adjacent sequences, e.g. the Kozak sequence. In cases where sequences encoding PKIN and its initiation codon and upstream regulatory sequences are inserted into the appropriate expression vector, no additional transcriptional or translational control signals may be needed. However, in cases where only coding sequence, or a fragment thereof, is inserted, exogenous translational control signals including an in-frame ATG initiation codon should be provided by the vector. Exogenous translational elements and initiation codons may be of various origins, both natural and synthetic. The e~ciency of expression may be enhanced by the inclusion of enhancers appropriate for the particular host cell system used. (See, e.g., Scharf, D.
et al. (1994) Results Probl.
Cell Differ. 20:125-I62.) Methods which are well known to those skilled in the art may be used to construct expression vectors containing sequences encoding PKIN and appropriate transcriptional and translational control elements. These methods include in vitro recombinant DNA techniques, synthetic techniques, and in vivo genetic recombination. (See, e.g., Sambrook, J. et al. (1989) Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Press, Plainview NY, ch. 4, 8, and 16-17; Ausubel, F.M. et al. (1995) Current Protocols in Molecular Biolo~y, John Wiley & Sons, New York NY, ch. 9, 13, and 16.) A variety of expression vector/host systems may be utilized to contain and express sequences encoding PKIN. These include, but are not limited to, microorganisms such as bacteria transformed with recombinant bacteriophage, plasmid, or cosmid DNA expression vectors;
yeast transformed with yeast expression vectors; insect cell systems infected with viral expression vectors (e.g., baculovirus);
plant cell systems transformed with viral expression vectors (e.g., cauliflower mosaic virus, CaMV, or tobacco mosaic virus, TMV) or with bacterial expression vectors (e.g., Ti or pBR322 plasmids); or animal cell systems. (See, e.g., Sambrook, supra; Ausubel, supra; Van Heeke, G. and S.M. Schuster (1989) J. Biol. Chem. 264:5503-5509; Engelhard, E.K. et al. (1994) Proc. Natl.
Acad. Sci. USA
91:3224-3227; Sandig, V. et al. (1996) Hum. Gene Ther. 7:1937-1945; Takamatsu, N. (1987) EMBO
J. 6:307-311; The McGraw Hill Yearbook of Science and Technolo~y (1992) McGraw Hill, New York NY, pp. 191-196; Logan, J. and T. Shenk (1984) Proc. Natl. Acad. Sci. USA
81:3655-3659; and Harrington, J.J. et al. (1997) Nat. Genet. 15:345-355.) Expression vectors derived from retroviruses, adenoviruses, or herpes or vaccinia viruses, or from various bacterial plasmids, may be used for delivery of nucleotide sequences to the targeted organ, tissue, or cell population. (See, e.g., Di Nicola, M. et al. (1998) Cancer Gen. Ther. 5(6):350-356; Yu, M. et al. (1993) Proc.
Natl. Acad. Sci. USA
90(13):6340-6344; Buller, R.M. et al. (1985) Nature 317(6040):813-815;
McGregor, D.P. et al. (1994) Mol. Immunol. 31(3):219-226; and Verma, LM. and N. Somia (1997) Nature 389:239-242.) The invention is not limited by the host cell employed.

In bacterial systems, a number of cloning and expression vectors may be selected depending upon the use intended for polynucleotide sequences encoding PKIN. For example, routine cloning, subcloning, and propagation of polynucleotide sequences encoding PKIN can be achieved using a multifunctional E. coli vector such as PBLUESCRIPT (Stratagene, La Jolla CA) or PSPORTl I plasmid (Life Technologies). Ligation of sequences encoding PI~IN into the vector's multiple cloning site disrupts the ZacZ gene, allowing a colorimetric screening procedure for identification of transformed bacteria containing recombinant molecules. In addition, these vectors may be useful for in vitro transcription, dideoxy sequencing, single strand rescue with helper phage, and creation of nested deletions in the cloned sequence. (See, e.g., Van Heeke, G. and S.M.
Schuster (1989) J. Biol.
Chem. 264:5503-5509.) When large quantities of PKIN are needed, e.g. for the production of antibodies, vectors which direct high level expression of PKIN may be used.
For example, vectors containing the strong, inducible SP6 or T7 bacteriophage promoter may be used.
Yeast expression systems may be used for production of PKIN. A number of vectors containing constitutive or inducible promoters, such as alpha factor, alcohol oxidase, and PGH
promoters, may be used in the yeast Saccharomyces cerevisiae or Pichia pastoris. In addition, such vectors direct either the secretion or intracellular retention of expressed proteins and enable integration of foreign sequences into the host genome for stable propagation. (See, e.g., Ausubel, 1995, supra;
Bitter, G.A. et al. (1987) Methods Enzymol. 153:516-544; and Scorer, C.A. et al. (1994) BiolTechnology 12:181-184.) Plant systems may also be used for expression of PKIN. Transcription of sequences encoding PI~IN may be driven by viral promoters, e.g., the 35S and 19S
promoters of CaMV used alone or in combination with the omega leader sequence from TMV (Takamatsu, N.
(1987) EMBO J.

6:307-311). Alternatively, plant promoters such as the small subunit of RUBISCO or heat shock promoters may be used. (See, e.g., Coruzzi, G. et al. (1984) EMBO J. 3:1671-1680; Brogue, R. et al.
(1984) Science 224:838-843; and Winter, J. et al. (1991) Results Probl. Cell Differ. 17:85-105.) These constructs can be introduced into plant cells by direct DNA transformation or pathogen-mediated transfection. (See, e.g., The McGraw Hill Yearbook of Science and Technolo~y (1992) McGraw Hill, New York NY, pp. 191-196.) In mammalian cells, a number of viral-based expression systems may be utilized. In cases where an adenovirus is used as an expression vector, sequences encoding PKIN
may be ligated into an adenovirus transcription/translation complex consisting of the late promoter and tripartite leader sequence. Insertion in a non-essential E1 or E3 region of the viral genome may be used to obtain infective virus which expresses PKIN in host cells. (See, e.g., Logan, J. and T. Shenk (1984) Proc.

Natl. Acad. Sci. USA 81:3655-3659.) In addition, transcription enhancers, such as the Rous sarcoma virus (RSV) enhancer, may be used to increase expression in mammalian host cells. SV40 or EBV-based vectors may also be used for high-level protein expression.
Human artificial chromosomes (HACs) may also be employed to deliver larger fragments of DNA than can be contained in and expressed from a plasmid. HACs of about 6 kb to 10 Mb are constructed and delivered via conventional delivery methods (liposomes, polycationic amino polymers, or vesicles) for therapeutic purposes. (See, e.g., Harrington, J.J. et al.
(1997) Nat. Genet. 15:345-355.) For long term production of recombinant proteins in mammalian systems, stable expression of PKIN in cell lines is preferred. For example, sequences encoding PI~IN can be transformed into cell lines using expression vectors which may contain viral origins of replication and/or endogenous expression elements and a selectable marker gene on the same or on a separate vector. Following the introduction of the vector, cells may be allowed to grow for about 1 to 2 days in enriched media before being switched to selective media. The purpose of the selectable marker is to confer resistance to a selective agent, and its presence allows growth and recovery of cells which successfully express the introduced sequences. Resistant clones of stably transformed cells may be propagated using tissue culture techniques appropriate to the cell type.
Any number of selection systems may be used to recover transformed cell lines.
These include, but are not limited to, the herpes simplex virus thymidine kinase and adenine phosphoribosyltransferase genes, for use in t7~ and apr cells, respectively.
(See, e.g., Wigler, M. et aI. (1977) Cell 11:223-232; Lowy, I. et al. (1980) Cell 22:817-823.) Also, antimetabolite, antibiotic, or herbicide resistance can be used as the basis for selection. For example, dlafr confers resistance to methotrexate; neo confers resistance to the aminoglycosides neomycin and G-418; and als and pat confer resistance to chlorsulfuron and phosphinotricin acetyltransferase, respectively. (See, e.g., Wigler, M. et al. (1980) Proc. Natl. Acad. Sci. USA 77:3567-3570; Colbere-Garapin, F. et al. (1981) J. Mol. Biol. 150:1-14.) Additional selectable genes have been described, e.g., trpB and hisD, which alter cellular requirements for metabolites. (See, e.g., Hartman, S.C. and R.C. Mulligan (1988) Proc.
Natl. Acad. Sci. USA 85:8047-8051.) Visible markers, e.g., anthocyanins, green fluorescent proteins (GFP; Clontech),13 glucuronidase and its substrate J3-glucuronide, or luciferase and its substrate luciferin may be used. These markers can be used not only to identify transformants, but also to quantify the amount of transient or stable protein expression attributable to a specific vector system.
(See, e.g., Rhodes, C.A. (1995) Methods Mol. Biol. 55:121-131.) Although the presence/absence of marker gene expression suggests that the gene of interest is also present, the presence and expression of the gene may need to be confirmed. For example, if the sequence encoding PKIN is inserted within a marker gene sequence, transformed cells containing sequences encoding PKIN can be identified by the absence of marker gene function. Alternatively, a marker gene can be placed in tandem with a sequence encoding PKIN under the control of a single promoter. Expression of the marker gene in response to induction or selection usually indicates expression of the tandem gene as well.
In general, host cells that contain the nucleic acid sequence encoding PKIN
and that express PKIN may be identified by a variety of procedures known to those of skill in the art. These procedures include, but are not limited to, DNA-DNA or DNA-RNA hybridizations, PCR
amplification, and protein bioassay or immunoassay techniques which include membrane, solution, or chip based technologies for the detection and/or quantification of nucleic acid or protein sequences.
Immunological methods for detecting and measuring the expression of PKIN using either specific polyclonal or monoclonal antibodies are known in the art. Examples of such techniques include enzyme-linked immunosorbent assays (ELISAs), radioimmunoassays (RIAs), and fluorescence activated cell sorting (FACS). A two-site, monoclonal-based immunoassay utilizing monoclonal antibodies reactive to two non-interfering epitopes on PI~IN is preferred, but a competitive binding assay may be employed. These and other assays are well known in the art. (See, e.g., Hampton, R. et al. (1990) Serological Methods, a Laboratory Manual, APS Press, St. Paul MN, Sect.
IV; Coligan, J.E. et al. (1997) Current Protocols in Immunolo~y, Greene Pub.
Associates and Wiley-Interscience, New York NY; and Pound, J.D. (1998) Immunochemical Protocols, Humana Press, Totowa NJ.) A wide variety of labels and conjugation techniques are known by those skilled in the art and may be used in various nucleic acid and amino acid assays. Means for producing labeled hybridization or PCR probes for detecting sequences related to polynucleotides encoding PKIN
include oligolabeling, nick translation, end-labeling, or PCR amplification using a labeled nucleotide.
Alternatively, the sequences encoding PKIN, or any fragments thereof, may be cloned into a vector for the production of an mRNA probe. Such vectors are known in the art, are commercially available, and may be used to synthesize RNA probes in vitro by addition of an appropriate RNA polymerase such as T7, T3, or SP6 and labeled nucleotides. These procedures may be conducted using a variety of commercially available kits, such as those provided by Amersham Pharmacia Biotech, Promega (Madison WI), and US Biochemical. Suitable reporter molecules or labels which may be used for ease of detection include radionuclides, enzymes, fluorescent, chemiluminescent, or chromogenic agents, as well as substrates, cofactors, inhibitors, magnetic particles, and the like.

Host cells transformed with nucleotide sequences encoding PKIN may be cultured under conditions suitable for the expression and recovery of the protein from cell culture. The protein produced by a transformed cell may be secreted or retained intracellularly depending on the sequence and/or the vector used. As will be understood by those of skill in the art, expression vectors containing polynucleotides which encode PKIN may be designed to contain signal sequences which direct secretion of PKIN through a prokaryotic or eukaryotic cell membrane.
In addition, a host cell strain may be chosen for its ability to modulate expression of the inserted sequences or to process the expressed protein in the desired fashion.
Such modifications of the polypeptide include, but are not limited to, acetylation, carboxylation, glycosylation, phosphorylation, lipidation, and acylation. Post-translational processing which cleaves a "prepro" or "pro" form of the protein may also be used to specify protein targeting, folding, and/or activity. Different host cells which have specific cellular machinery and characteristic mechanisms for post-translational activities (e.g., CHO, HeLa, MDCK, HEK293, and WI38) are available from the American Type Culture Collection (ATCC, Manassas VA) and may be chosen to ensure the correct modification and processing of the foreign protein.
In another embodiment of the invention, natural, modified, or recombinant nucleic acid sequences encoding PKIN may be ligated to a heterologous sequence resulting in translation of a fusion protein in any of the aforementioned host systems. For example,, a chimeric PKIN protein containing a heterologous moiety that can be recognized by a commercially available antibody may facilitate the screening of peptide libraries for inhibitors of PKIN activity.
Heterologous protein and peptide moieties may also facilitate purification of fusion proteins using commercially available affinity matrices. Such moieties include, but are not limited to, glutathione S-transferase (GST), maltose binding protein (MBP), thioredoxin (Trx), calmodulin binding peptide (CBP), 6-His, FLAG, c-myc, and hemagglutinin (HA). GST, MBP, Trx, CBP, and 6-His enable purification of their cognate fusion proteins on immobilized glutathione, maltose, phenylarsine oxide, calmodulin, and metal-chelate resins, respectively. FLAG, c-nayc, and hemagglutinin (HA) enable immunoaffinity purification of fusion proteins using commercially available monoclonal and polyclonal antibodies that specifically recognize these epitope tags. A fusion protein may also be engineered to contain a proteolytic cleavage site located between the PKIN encoding sequence and the heterologous protein sequence, so that PKIN
may be cleaved away from the heterologous moiety following purification.
Methods for fusion protein expression and purification are discussed in Ausubel (1995, su ra, ch. 10). A
variety of commercially available kits may also be used to facilitate expression and purification of fusion proteins.
In a further embodiment of the invention, synthesis of radiolabeled PKIN may be achieved in vitro using the TNT rabbit reticulocyte lysate or wheat germ extract system (Promega). These systems couple transcription and translation of protein-coding sequences operably associated with the T7, T3, or SP6 promoters. Translation takes place in the presence of a radiolabeled amino acid precursor, for example, 35S-methionine.
PKIN of the present invention or fragments thereof may be used to screen for compounds that specifically bind to PKIN. At least one and up to a plurality of test compounds may be screened for specific binding to PKIN. Examples of test compounds include antibodies, oligonucleotides, proteins (e.g., receptors), or small molecules.
In one embodiment, the compound thus identified is closely related to the natural ligand of l0 PKIN, e.g., a ligand or fragment thereof, a natural substrate, a structural or functional mimetic, or a natural binding partner. (See, e.g., Coligan, J.E. et al. (1991) Current Protocols in Immunolo~y 1(2):
Chapter 5.) Similarly, the compound can be closely related to the natural receptor to which PKIN
binds, or to at least a fragment of the receptor, e.g., the ligand binding site. In either case, the compound can be rationally designed using known techniques. In one embodiment, screening for these compounds involves producing appropriate cells which express PKIN, either as a secreted protein or on the cell membrane. Preferred cells include cells from mammals, yeast, Drosophila, or E.
coli. Cells expressing PKIN or cell membrane fractions which contain PKIN are then contacted with a test compound and binding, stimulation, or inhibition of activity of either PKIN or the compound is analyzed. .
An assay may simply test binding of a test compound to the polypeptide, wherein binding is detected by a fluorophore, radioisotope, enzyme conjugate, or other detectable label. For example, the assay may comprise the steps of combining at least one test compound with PKIN, either in solution or affixed to a solid support, and detecting the binding of PKIN to the compound. Alternatively, the assay may detect or measure binding of a test compound in the presence of a labeled competitor.
Additionally, the assay may be carried out using cell-free preparations, chemical libraries, or natural product mixtures, and the test compounds) may be free in solution or affixed to a solid support.
PKIN of the present invention or fragments thereof may be used to screen for compounds that modulate the activity of PKIN. Such compounds may include agonists, antagonists, or partial or inverse agonists. In one embodiment, an assay is performed under conditions permissive for PKIN
activity, wherein PKIN is combined with at least one test compound, and the activity of PKIN in the presence of a test compound is compared with the activity of PKIN in the absence of the test compound. A change in the activity of PKIN in the presence of the test compound is indicative of a compound that modulates the activity of PKIN. Alternatively, a test compound is combined with an in vitro or cell-free system comprising PKIN under conditions suitable for PKIN
activity, and the assay is performed. In either of these assays, a test compound which modulates the activity of PKIN may do so indirectly and need not come in direct contact with the test compound.
At least one and up to a plurality of test compounds may be screened.
In another embodiment, polynucleotides encoding PKIN or their mammalian homologs may be "knocked out" in an animal model system using homologous recombination in embryonic stem (ES) cells. Such techniques are well known in the art and are useful for the generation of animal models of human disease. (See, e.g., U.S. Patent Number 5,175,383 and U.S. Patent Number 5,767,337.) For example, mouse ES cells, such as the mouse 129/SvJ cell line, are derived from the early mouse embryo and grown in culture. The ES cells are transformed with a vector containing the gene of interest disrupted by a marker gene, e.g., the neomycin phosphotransferase gene (neo; Capecchi, M.R. (1989) Science 244:1288-1292). The vector integrates into the corresponding region of the host genome by homologous recombination. Alternatively, homologous recombination takes place using the Cre-loxP system to knockout a gene of interest in a tissue- or developmental stage-specific manner (Maxth, J.D. (1996) Clin. Invest. 97:1999-2002; Wagner, K.U. et al. (1997) Nucleic Acids Res.
25:4323-4330). Transformed ES cells are identified and microinjected into mouse cell blastocysts such as those from the C57BL/6 mouse strain. The blastocysts are surgically transferred to pseudopregnant dams, and the resulting chimeric progeny are genotyped and bred to produce heterozygous or homozygous strains. Transgenic animals thus generated may be tested with potential therapeutic or toxic agents.
Polynucleotides encoding PKIN may also be manipulated in vitro in ES cells derived from human blastocysts. Human ES cells have the potential to differentiate into at least eight separate cell lineages including endoderm, mesoderm, and ectodermal cell types. These cell lineages differentiate into, for example, neural cells, hematopoietic lineages, and cardiomyocytes (Thomson, J.A. et al.
(1998) Science 282:1145-1147).
Polynucleotides encoding PKIN can also be used to create "knockin" humanized animals (pigs) or transgenic animals (mice or rats) to model human disease. With knockin technology, a region of a polynucleotide encoding PKIN is injected into animal ES cells, and the injected sequence integrates into the animal cell genome. Transformed cells are injected into blastulae, and the blastulae are implanted as described above. Transgenic progeny or inbred lines are studied and treated with potential pharmaceutical agents to obtain information on treatment of a human disease. Alternatively, a mammal inbred to overexpress PKIN, e.g., by secreting PKIN in its milk, may also serve as a convenient source of that protein (Janne, J. et al. (1998) Biotechnol. Annu.
Rev. 4:55-74).

THERAPEUTICS
Chemical and structural similarity, e.g., in the context of sequences and motifs, exists between regions of PKIN and human kinases. In addition, the expression of PKIN is closely associated with bladder cancer, prostatic, ovarian, brain, colon, ileum, penis, skin, adrenal tumor, digestive, and cancerous tissues. Therefore, PI~IN appears to play a role in cancer, immune disorders, disorders affecting growth and development, cardiovascular diseases, and lipid disorders. In the treatment of disorders associated with increased PI~IN expression or activity, it is desirable to decrease the expression or activity of PKIN. In the treatment of disorders associated with decreased PI~IN
expression or activity, it is desirable to increase the expression or activity of PI~IN.
Therefore, in one embodiment, PKIN or a fragment or derivative thereof may be administered to a subject to treat or prevent a disorder associated with decreased expression or activity of PI~IN.
Examples of such disorders include, but are not limited to, a cancer, such as adenocarcinoma, leukemia, lymphoma, melanoma, myeloma, sarcoma, teratocarcinoma, and, in particular, cancers of the adrenal gland, bladder, bone, bone marrow, brain, breast, cervix, gall bladder, ganglia, gastrointestinal tract, heart, kidney, liver, lung, muscle, ovary, pancreas, parathyroid, penis, prostate, salivary glands, skin, spleen, testis, thymus, thyroid, and uterus, leukemias such as multiple myeloma and lymphomas such as Hodgkin's disease; an immune disorder, such as acquired immunodeficiency syndrome (AIDS), Addison's disease, adult respiratory distress syndrome, allergies, ankylosing spondylitis, amyloidosis, anemia, asthma, atherosclerosis, autoimmune hemolytic anemia, autoimmune thyroiditis, autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED), bronchitis, cholecystitis, contact dermatitis, Crohn's disease, atopic dermatitis, dermatomyositis, diabetes mellitus, emphysema, episodic lymphopenia with lymphocytotoxins, erythroblastosis fetalis, erythema nodosum, atrophic gastritis, glomerulonephritis, Goodpasture's syndrome, gout, Graves' disease, Hashimoto's thyroiditis, hypereosinophilia, irritable bowel syndrome, multiple sclerosis, myasthenia gravis, myocardial or pericardial inflammation, osteoarthritis, osteoporosis, pancreatitis, polymyositis, psoriasis, Reiter's syndrome, rheumatoid arthritis, scleroderma, Sjogren's syndrome, systemic anaphylaxis, systemic lupus erythematosus, systemic sclerosis, thrombocytopenic purpura, ulcerative colitis, uveitis, Werner syndrome, complications of cancer, hemodialysis, and extracorporeal circulation, viral, bacterial, fungal, parasitic, protozoal, and hehninthic infections, and trauma; a growth and developmental disorder, such as actinic keratosis, arteriosclerosis, atherosclerosis, bursitis, cirrhosis, hepatitis, mixed connective tissue disease (MCTD), myelofibrosis, paroxysmal nocturnal hemoglobinuria, polycythemia vera, psoriasis, primary thrombocythemia, and cancers including adenocarcinoma, leukemia, lymphoma, melanoma, myeloma, sarcoma, teratocarcinoma, and, in particular, cancers of the adrenal gland, bladder, bone, bone marrow, brain, breast, cervix, gall bladder, ganglia, gastrointestinal tract, heaxt, kidney, liver, lung, muscle, ovary, pancreas, parathyroid, penis, prostate, salivary glands, skin, spleen, testis, thymus, thyroid, and uterus, renal tubular acidosis, anemia, Cushing's syndrome, achondroplastic dwarfism, Duchenne and Becker muscular dystrophy, epilepsy, gonadal dysgenesis, WAGR syndrome (Wilins' tumor, aniridia, genitourinary abnormalities, and mental retardation), Smith-Magenis syndrome, myelodysplastic syndrome, hereditary mucoepithelial dysplasia, hereditary keratodermas, hereditary neuropathies such as Charcot-Marie-Tooth disease and neurofibromatosis, hypothyroidism, hydrocephalus, seizure disorders such as Syndenham's chorea and cerebral palsy, spina bifida, anencephaly, craniorachischisis, congenital glaucoma, cataract, and sensorineural hearing loss; a cardiovascular disease, such as arteriovenous fistula, atherosclerosis, hypertension, vasculitis, Raynaud's disease, aneurysms, arterial dissections, varicose veins, thrombophlebitis and phlebothrombosis, vascular tumors, and complications of thrombolysis, balloon angioplasty, vascular replacement, and coronary artery bypass graft surgery, congestive heart failure, ischemic heaxt disease, angina pectoris, myocardial infarction, hypertensive heart disease, degenerative valvular heart disease, calcific aortic valve stenosis, congenitally bicuspid aortic valve, mitral annular calcification, mitral valve prolapse, rheumatic fever and rheumatic heart disease, infective endocarditis, nonbacterial thrombotic endocarditis, endocarditis of systemic lupus erythematosus, carcinoid heart disease, cardiomyopathy, myocarditis, pericarditis, neoplastic heart disease, congenital heart disease, and complications of cardiac transplantation, congenital lung anomalies, atelectasis, pulmonary congestion and edema, pulmonary embolism, pulmonary hemorrhage, pulmonary infarction, pulmonary hypertension, vascular sclerosis, obstructive pulmonary disease, restrictive pulmonary disease, chronic obstructive pulmonary disease, emphysema, chronic bronchitis, bronchial asthma, bronchiectasis, bacterial pneumonia, viral and mycoplasmal pneumonia, lung abscess, pulmonary tuberculosis, diffuse interstitial diseases, pneumoconioses, sarcoidosis, idiopathic pulmonary fibrosis, desquamative interstitial pneumonitis, hypersensitivity pneumonitis, pulmonary eosinophilia bronchiolitis obliterans-organizing pneumonia, diffuse pulmonary hemorrhage syndromes, Goodpasture's syndromes, idiopathic pulmonary hemosiderosis, pulmonary involvement in collagen-vascular disorders, pulmonary alveolar proteinosis, lung tumors, inflammatory and noninflammatory pleural effusions, pneumothorax, pleural tumors, drug-induced lung disease, radiation-induced lung disease, and complications of lung transplantation; and a lipid disorder such as fatty liver, cholestasis, primary biliary cirrhosis, caxnitine deficiency, caxnitine palmitoyltransferase deficiency, myoadenylate deaminase deftciency, hypertriglyceridemia, lipid storage disorders such Fabry's disease, Gaucher's disease, Niemann-Pick's disease, metachromatic leukodystrophy, adrenoleukodystrophy, GMZ gangliosidosis, and ceroid lipofuscinosis, abetalipoproteinemia, Tangier disease, hyperlipoproteinemia, diabetes mellitus, lipodystrophy, lipomatoses, acute panniculitis, disseminated fat necrosis, adiposis dolorosa, lipoid adrenal hyperplasia, minimal change disease, lipomas, atherosclerosis, hypercholesterolemia, hypercholesterolemia with hypertriglyceridemia, primary hypoalphalipoproteinemia, hypothyroidism, renal disease, liver disease, lecithin:cholesterol acyltransferase deficiency, cerebrotendinous xanthomatosis, sitosterolemia, hypocholesterolemia, Tay-Sachs disease, Sandhoff's disease, hyperlipidemia, hyperlipemia, lipid myopathies, and obesity.
In another embodiment, a vector capable of expressing PKIN or a fragment or derivative thereof may be administered to a subject to treat or prevent a disorder associated with decreased expression or activity of PKIN including, but not limited to, those described above.
In a further embodiment, a composition comprising a substantially purified PKTN in conjunction with a suitable pharmaceutical carrier may be administered to a subject to treat or prevent a disorder associated with decreased expression or activity of PKIN including, but not limited to, those provided above.
In still another embodiment, an agonist which modulates the activity of PKIN
may be administered to a subject to treat or prevent a disorder associated with decreased expression or activity of PKIN including, but not limited to, those listed above.
In a further embodiment, an antagonist of PKIN may be administered to a subject to treat or prevent a disorder associated with increased expression or activity of PKIN.
Examples of such disorders include, but are not limited to, those cancer, immune disorders, disorders affecting growth and development, cardiovascular diseases, and lipid disorders described above.
In one aspect, an antibody which specifically binds PKIN may be used directly as an antagonist or indirectly as a targeting or delivery mechanism for bringing a pharmaceutical agent to cells or tissues which express PKIN.
In an additional embodiment, a vector expressing the complement of the polynucleotide encoding PKIN may be administered to a subject to treat or prevent a disorder associated with increased expression or activity of PKIN including, but not limited to, those described above.
In other embodiments, any of the proteins, antagonists, antibodies, agonists, complementary sequences, or vectors of the invention may be administered in combination with other appropriate therapeutic agents. Selection of the appropriate agents for use in combination therapy may be made by one of ordinary skill in the art, according to conventional pharmaceutical principles. The combination of therapeutic agents may act synergistically to effect the treatment or prevention of the various disorders described above. Using this approach, one may be able to achieve therapeutic SO

efficacy with lower dosages of each agent, thus reducing the potential for adverse side effects.
An antagonist of PKIN may be produced using methods which are generally known in the art.
In particular, purified PKIN may be used to produce antibodies or to screen libraries of pharmaceutical agents to identify those which specifically bind PI~IN.
Antibodies to PKIN may also be generated using methods that are well known in the art. Such antibodies may include, but are not limited to, polyclonal, monoclonal, chimeric, and single chain antibodies, Fab fragments, and fragments produced by a Fab expression library. Neutralizing antibodies (i.e., those which inhibit dimer formation) are generally preferred for therapeutic use.
For the production of antibodies, various hosts including goats, rabbits, rats, mice, humans, and others may be immunized by injection with PKIN or with any fragment or oligopeptide thereof which has immunogenic properties. Depending on the host species, various adjuvants may be used to increase immunological response. Such adjuvants include, but are not limited to, Freund's, mineral gels such as aluminum hydroxide, and surface active substances such as lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, KLH, and dinitrophenol. Among adjuvants used in humans, BCG
(bacilli Calinette-Guerin) and Corynebacterium parvum are especially preferable.
It is preferred that the oligopeptides, peptides, or fragments used to induce antibodies to PI~IN
have an amino acid sequence consisting of at least about 5 amino acids, and generally will consist of at least about 10 anuno acids. It is also preferable that these oligopeptides, peptides, or fragments are identical to a portion of the amino acid sequence of the natural protein.
Short stretches of PI~IN
amino acids may be fused with those of another protein, such as I~LH, and antibodies to the chimeric molecule may be produced.
Monoclonal antibodies to PKIN may be prepared using any technique which provides for the production of antibody molecules by continuous cell lines in culture. These include, but are not limited to, the hybridoma technique, the human B-cell hybridoma technique, and the EB
V-hybridoma technique. (See, e.g., I~ohler, G. et al. (1975) Nature 256:495-497; Kozbor, D. et al. (1985) J.
hnmunol. Methods 81:31-42; Cote, R.J. et al. (1983) Proc. Natl. Acad. Sci. USA
80:2026-2030; and Cole, S.P. et al. (1984) Mol. Cell Biol. 62:109-120.) In addition, techniques developed for the production of "chimeric antibodies,"
such as the splicing of mouse antibody genes to human antibody genes to obtain a molecule with appropriate antigen specificity and biological activity, can be used. (See, e.g., Morrison, S.L. et al. (1984) Proc.
Natl. Acad. Sci. USA 81:6851-6855; Neuberger, M.S. et al. (1984) Nature 312:604-608; and Takeda, S. et al. (1985) Nature 314:452-454.) Alternatively, techniques described for the production of single chain antibodies may be adapted, using methods known in the art, to produce PI~IN-specific single chain antibodies. Antibodies with related specificity, but of distinct idiotypic composition, may be generated by chain shuffling from random combinatorial immunoglobulin libraries. (See, e.g., Burton, D.R. (1991) Proc. Natl. Acad. Sci. USA 88:10134-10137.) Antibodies may also be produced by inducing in vivo production in the lymphocyte population or by screening immunoglobulin libraries or panels of highly specific binding reagents as disclosed in the literature. (See, e.g., Orlandi, R. et al. (1989) Proc. Natl. Acad. Sci.
USA 86:3833-3837; Winter, G. et al. (1991) Nature 349:293-299.) Antibody fragments which contain specific binding sites for PKIN may also be generated.
For example, such fragments include, but are not limited to, F(ab~2 fragments produced by pepsin digestion of the antibody molecule and Fab fragments generated by reducing the disulfide bridges of the F(ab~2 fragments. Alternatively, Fab expression libraries may be constructed to allow rapid and easy identification of monoclonal Fab fragments with the desired specificity.
(See, e.g., Huse, W.D.
et al. (1989) Science 246:1275-1281.) Various immunoassays may be used for screening to identify antibodies having the desired specificity. Numerous protocols for competitive binding or immunoradiometric assays using either polyclonal or monoclonal antibodies with established specificities are well known in the art. Such immunoassays typically involve the measurement of complex formation between PKIN and its specific antibody. A two-site, monoclonal-based immunoassay utilizing monoclonal antibodies reactive to two non-interfering PKIN epitopes is generally used, but a competitive binding assay may also be 2o employed (Pound, supra).
Various methods such as Scatchard analysis in conjunction with radioimmunoassay techniques may be used to assess the affinity of antibodies for PI~IN. Affinity is expressed as an association constant, I~, which is defined as the molar concentration of PKIN-antibody complex divided by the molar concentrations of free antigen and free antibody under equilibrium conditions. The I~
determined for a preparation of polyclonal antibodies, which are heterogeneous in their affinities for multiple PI~iN epitopes, represents the average affinity, or avidity, of the antibodies for PKIN. The Ka determined for a preparation of monoclonal antibodies, which are monospecific for a particular PKIN epitope, represents a true measure of affinity. High-affinity antibody preparations with I~
ranging from about 109 to 1012 Llmole are preferred for use in immunoassays in which the PI~IN-antibody complex must withstand rigorous manipulations. Low-affinity antibody preparations with I~
ranging from about 106 to 10' L/mole are preferred for use in immunopurification and similar procedures which ultimately require dissociation of PKIN, preferably in active form, from the antibody (Catty, D. (1988) Antibodies, Volume I: A Practical Approach, IRL Press, Washington DC; Liddell, J.E. and A. Cryer (1991) A Practical Guide to Monoclonal Antibodies, John Wiley & Sons, New York NY).
The titer and avidity of polyclonal antibody preparations may be further evaluated to determine the quality and suitability of such preparations for certain downstream applications. For example, a polyclonal antibody preparation containing at least 1-2 mg specific antibody/ml, preferably 5-10 mg specific antibodylml, is generally employed in procedures requiring precipitation of PI~IN-antibody complexes. Procedures for evaluating antibody specificity, titer, and avidity, and guidelines for antibody quality and usage in various applications, are generally available.
(See, e.g., Catty, supra, and Coligan et al. s-upra.) In another embodiment of the invention, the polynucleotides encoding PKIN, or any fragment or complement thereof, may be used for therapeutic purposes. In one aspect, modifications of gene expression can be achieved by designing complementary sequences or antisense molecules (DNA, RNA, PNA, or modified oligonucleotides) to the coding or regulatory regions of the gene encoding PI~IN. Such technology is well known in the art, and antisense oligonucleotides or larger fragments can be designed from various locations along the coding or control regions of sequences encoding PKIN. (See, e.g., Agrawal, S., ed. (1996) Antisense Therapeutics, Humana Press Inc., Totawa NJ.) In therapeutic use, any gene delivery system suitable for introduction of the antisense sequences into appropriate target cells can be used. Antisense sequences can be delivered intracellularly in the form of.an expression plasmid which, upon transcription, produces a sequence complementary to at least a portion of the cellular sequence encoding the target protein. (See, e.g., Slater, J.E. et al. (1998) J. Allergy Clin. Immunol. 102(3):469-475; and Scanlon, K.J. et al. (1995) 9(13):1288-1296.) Antisense sequences can also be introduced intracellularly through the use of viral vectors, such as retrovirus and adeno-associated virus vectors. (See, e.g., Miller, A.D. (1990) Blood 76:271; Ausubel, supra; Uckert, W. and W. Walther (1994) Pharmacol. Ther.
63(3):323-347.) Other gene delivery mechanisms include liposome-derived systems, artificial viral envelopes, and other systems known in the art. (See, e.g., Rossi, J.J. (1995) Br. Med. Bull.
51(1):217-225; Boado, R.J. et al. (1998) J. Pharm. Sci. 87(11):1308-1315; and Morris, M.C. et al. (1997) Nucleic Acids Res.
25(14):2730-2736.) In another embodiment of the invention, polynucleotides encoding PHIN may be used for somatic or gerxnline gene therapy. Gene therapy may be performed to (i) correct a genetic deficiency (e.g., in the cases of severe combined immunodeficiency (SCID)-Xl disease characterized by X-linked inheritance (Cavazzana-Calvo, M. et al. (2000) Science 288:669-672), severe combined immunodeficiency syndrome associated with an inherited adenosine deaminase (ADA) deficiency (Blaese, R.M. et al. (1995) Science 270:475-480; Bordignon, C. et al. (1995) Science 270:470-475), cystic fibrosis (Zabner, J. et al. (1993) Cell 75:207-216; Crystal, R.G. et al. (1995) Hum. Gene Therapy 6:643-666; Crystal, R.G. et al. (1995) Hum. Gene Therapy 6:667-703), thalassamias, familial hypercholesterolemia, and hemophilia resulting from Factor VIII or Factor IX
deficiencies (Crystal, R.G. (1995) Science 270:404-410; Verma, LM. and N. Somia (1997) Nature 389:239-242)), (ii) express a conditionally lethal gene product (e.g., in the case of cancers which result from unregulated cell proliferation), or (iii) express a protein which affords protection against intracellular parasites (e.g., against human retroviruses, such as human immunodeficiency virus (HIV) (Baltimore, D. (1988) Nature 335:395-396; Poeschla, E. et al. (1996) Proc. Natl. Acad. Sci. USA.
93:11395-11399), hepatitis B or C virus (HBV, HCV); fungal parasites, such as Candida albicans and Paracoccidioides brasiliensis; and protozoan parasites such as Plasmodium falciparum and Trypanosoma cruzi). In the case where a genetic deficiency in PKIN expression or regulation causes disease, the expression of PKIN from an appropriate population of transduced cells may alleviate the clinical manifestations caused by the genetic deficiency.
In a further embodiment of the invention, diseases or disorders caused by deficiencies in PKIN are treated by constructing mammalian expression vectors encoding PKIN
and introducing these vectors by mechanical means into PKIN-deficient cells. Mechanical transfer technologies for use with cells in vivo or ex vitro include (i) direct DNA microinjection into individual cells, (ii) ballistic gold particle delivery, (iii) liposome-mediated transfection, (iv) receptor-mediated gene transfer, and (v) the use of DNA transposons (Morgan, R.A. and W.F. Anderson (1993) Annu.
Rev. Biochem.
62:191-217; Ivics, Z. (1997) Cell 91:501-510; Boulay, J-L. and H. Recipon (1998) Curr. Opin.
Biotechnol. 9:445-450).
Expression vectors that may be effective for the expression of PKIN include, but are not limited to, the PCDNA 3.1, EPITAG, PRCCMV2, PREP, PVAX vectors (Invitrogen, Carlsbad CA), PCMV-SCRIPT, PCMV-TAG, PEGSH/PERV (Stratagene, La Jolla CA), and PTET-OFF, PTET-ON, PTRE2, PTRE2-LUC, PTK-HYG (Clontech, Palo Alto CA). PKIN may be expressed using (i) a constitutively active promoter, (e.g., from cytomegalovirus (CMV), Rous sarcoma virus (RSV), SV40 virus, thymidine kinase (TK), or ~3-actin genes), (ii) an inducible promoter (e.g., the tetracycline-regulated promoter (Gossen, M. and H. Bujard (1992) Proc. Natl.
Acad. Sci. USA
89:5547-5551; Gossen, M. et al. (1995) Science 268:1766-1769; Rossi, F.M.V.
and H.M. Blau (1998) Curr. Opin. Biotechnol. 9:451-456), commercially available in the T-REX
plasmid (Invitxogen)); the ecdysone-inducible promoter (available in the plasmids PVGRXR and PIND;
Invitrogen); the FK506/rapamycin inducible promoter; or the RU486/mifepristone inducible promoter (Rossi, F.M.V.

and Blau, H.M. supra)), or (iii) a tissue-specific promoter or the native promoter of the endogenous gene encoding PKIN from a normal individual.
Commercially available liposome transformation kits (e.g., the PERFECT LIPID
TRANSFECTION KIT, available from Invitrogen) allow one with ordinary skill in the art to deliver polynucleotides to target cells in culture and require minimal effort to optimize experimental parameters. In the alternative, transformation is performed using the calcium phosphate method (Graham, F.L. and A.J. Eb (1973) Virology 52:456-467), or by electroporation (Neumann, E. et al.
(1982) EMBO J. 1:841-845). The introduction of DNA to primary cells requires modification of these standardized mammalian transfection protocols.
In another embodiment of the invention, diseases or disorders caused by genetic defects with respect to PKIN expression are treated by constructing a retrovirus vector consisting of (i) the polynucleotide encoding PKIN under the control of an independent promoter or the retrovirus long terminal repeat (LTR) promoter, (ii) appropriate RNA packaging signals, and (iii) a Rev-responsive element (RRE) along with additional retrovirus ciS-acting RNA sequences and coding sequences required for efficient vector propagation. Retrovirus vectors (e.g., PFB and PFBNEO) are commercially available (Stratagene) and are based on published data (Riviere, I. et al. (1995) Proc.
Natl. Acad. Sci. USA 92:6733-6737), incorporated by reference herein. The vector is propagated in an appropriate vector producing cell line (VPCL) that expresses an envelope gene with a tropism for receptors on the target cells or a promiscuous envelope protein such as VSVg (Armentano, D. et al.
(1987) J. Virol. 61:1647-1650; Bender, M.A. et al. (1987) J. Virol. 61:1639-1646; Adam, M.A. and A.D. Miller (1988) J. Virol. 62:3802-3806; Dull, T. et al. (1998) J. Virol.
72:8463-8471; Zufferey, R. et al. (1998) J. Virol. 72:9873-9880). U.S. Patent Number 5,910,434 to Rigg ("Method for obtaining retrovirus packaging cell lines producing high transducing efficiency retroviral supernatant") discloses a method for obtaining retrovirus packaging cell lines and is hereby incorporated by reference.
Propagation of retrovirus vectors, transduction of a populafiion of cells (e.g., CD4+ T-cells), and the return of transduced cells to a patient are procedures well known to persons skilled in the art of gene therapy and have been well documented (Ranga, U. et al. (1997) J. Virol.
71:7020-7029; Bauer, G. et al. (1997) Blood 89:2259-2267; Bonyhadi, M.L. (1997) J. Virol. 71:4707-4716;
Ranga, U. et al. (1998) Proc. Natl. Acad. Sci. USA 95:1201-1206; Su, L. (1997) Blood 89:2283-2290).
In the alternative, an adenovirus-based gene therapy delivery system is used to deliver polynucleotides encoding PKIN to cells which have one or more genetic abnormalities with respect to the expression of PKIN. The construction and packaging of adenovirus-based vectors are well known to those with ordinary skill in the art. Replication defective adenovirus vectors have proven to be versatile for importing genes encoding immunoregulatory proteins into intact islets in the pancreas (Csete, M.E. et al. (1995) Transplantation 27:263-268). Potentially useful adenoviral vectors are described in U.S. Patent Number 5,707,618 to Armentano ("Adenovirus vectors for gene therapy"), hereby incorporated by reference. For adenoviral vectors, see also Antinozzi, P.A. et al. (1999) Annu. Rev. Nutr. 19:511-544 and Verma, LM. and N. Somia (1997) Nature 18:389:239-242, both incorporated by reference herein.
In another alternative, a herpes-based, gene therapy delivery system is used to deliver polynucleotides encoding PKIN to target cells which have one or more genetic abnormalities with respect to the expression of PKIN. The use of herpes simplex virus (HSV)-based vectors may be especially valuable for introducing PKIN to cells of the central nervous system, for which HSV has a tropism. The construction and packaging of herpes-based vectors are well known to those with ordinary skill in the art. A replication-competent herpes simplex virus (HSV) type 1-based vector has been used to deliver a reporter gene to the eyes of primates (Liu, X. et al.
(1999) Exp. Eye Res.
169:385-395). The construction of a HSV-1 virus vector has also been disclosed in detail in U.S.
Patent Number 5,804,413 to DeLuca ("Herpes simplex virus strains for gene transfer"), which is hereby incorporated by reference. U.S. Patent Number 5,804,413 teaches the use of recombinant HSV d92 which consists of a genome containing at least one exogenous gene to be transferred to a cell under the control of the appropriate promoter for purposes including human gene therapy. Also taught by this patent are the construction and use of recombinant HSV strains deleted for ICP4, ICP27 and ICP22. For HSV vectors, see also Goins, W.F. et al. (1999) J. Virol.
73:519-532 and Xu, H. et al. (1994) Dev. Biol. 163:152-161, hereby incorporated by reference. The manipulation of cloned herpesvirus sequences, the generation of recombinant virus following the transfection of multiple plasmids containing different segments of the large herpesvirus genomes, the growth and propagation of herpesvirus, and the infection of cells with herpesvirus are techniques well known to those of ordinary skill in the art.
In another alternative, an alphavirus (positive, single-stranded RNA virus) vector is used to deliver polynucleotides encoding PKIN to target cells. The biology of the prototypic alphavirus, Semliki Forest Virus (SFV), has been studied extensively and gene transfer vectors have been based on the SFV genome (Garoff, H. and K.-J. Li (1998) Curr. Opin. Biotechnol.
9:464-469). During alphavirus RNA replication, a subgenomic RNA is generated that normally encodes the viral capsid proteins. This subgenomic RNA replicates to higher levels than the full length genomic RNA, resulting in the overproduction of capsid proteins relative to the viral proteins with enzymatic activity (e.g., protease and polymerase). Similarly, inserting the coding sequence for PKIN into the alphavirus genome in place of the capsid-coding region results in the production of a large number of PKIN-coding RNAs and the synthesis of high levels of PKIN in vector transduced cells. While alphavirus infection is typically associated with cell lysis within a few days, the ability to establish a persistent infection in hamster normal kidney cells (BHK-21) with a variant of Sindbis virus (SIN) indicates that the lytic replication of alphaviruses can be altered to suit the needs of the gene therapy application (Dryga, S.A. et al. (1997) Virology 228:74-83). The wide host range of alphaviruses will allow the introduction of PKIN into a variety of cell types. The specific transduction of a subset of cells in a population may require the sorting of cells prior to transduction. The methods of manipulating infectious cDNA clones of alphaviruses, performing alphavirus cDNA and RNA
transfections, and performing alphavirus infections, are well known to those with ordinary skill in the art.
Oligonucleotides derived from the transcription initiation site, e.g., between about positions -10 and +10 from the start site, may also be employed to inhibit gene expression.
Similarly, inhibition can be achieved using triple helix base-pairing methodology. Triple helix pairing is useful because it causes inhibition of the ability of the double helix to open sufficiently for the binding of polymerases, transcription factors, or regulatory molecules. Recent therapeutic advances using triplex DNA have been described in the literature. (See, e.g., Gee, J.E. et al. (1994) in Huber, B.E. and B.I. Carr, Molecular and Immunolo~ic Approaches, Futura Publishing, Mt. Kisco NY, pp. 163-177.) A
complementary sequence or antisense molecule may also be designed to block translation of mRNA
by preventing the transcript from binding to ribosomes.
Ribozymes, enzymatic RNA molecules, may also be used to catalyze the specific cleavage of RNA. The mechanism of ribozyme action involves sequence-specific hybridization of the ribozyme molecule to complementary target RNA, followed by endonucleolytic cleavage.
For example, engineered hammerhead motif ribozyme molecules may specifically and efficiently catalyze endonucleolytic cleavage of sequences encoding PKIN.
Specific ribozyme cleavage sites within any potential RNA target are initially identified by scanning the target molecule for ribozyme cleavage sites, including the following sequences: GUA, GUU, and GUC. Once identified, short RNA sequences of between 15 and 20 ribonucleotides, corresponding to the region of the target gene containing the cleavage site, may be evaluated for secondary structural features which may render the oligonucleotide inoperable.
The suitability of candidate targets may also be evaluated by testing accessibility to hybridization with complementary oligonucleotides using ribonuclease protection assays.
Complementary ribonucleic acid molecules and ribozymes of the invention may be prepared by any method known in the art for the synthesis of nucleic acid molecules.
These include techniques for chemically synthesizing oligonucleotides such as solid phase phosphoramidite chemical synthesis.
Alternatively, RNA molecules may be generated by in vitro and in vivo transcription of DNA
sequences encoding PKIN. Such DNA sequences may be incorporated into a wide variety of vectors with suitable RNA polymerase promoters such as T7 or SP6. Alternatively, these cDNA constructs that synthesize complementary RNA, constitutively or inducibly, can be introduced into cell lines, cells, or tissues.
RNA molecules may be modified to increase intracellular stability and half life. Possible modifications include, but are not limited to, the addition of flanking sequences at the 5' and/or 3' ends of the molecule, or the use of phosphorothioate or 2' O-methyl rather than phosphodiesterase linkages within the backbone of the molecule. This concept is inherent in the production of PNAs and can be extended in all of these molecules by the inclusion of nontraditional bases such as inosine, queosine, and wybutosine, as well as acetyl-, methyl-, thio-, and similarly modified forms of adenine, cytidine, guanine, thymine, and uridine which are not as easily recognized by endogenous endonucleases.
An additional embodiment of the invention encompasses a method for screening for a compound which is effective in altering expression of a polynucleotide encoding PKIN. Compounds which may be effective in altering expression of a specific polynucleotide may include, but are not limited to, oligonucleotides, antisense oligonucleotides, triple helix-forming oligonucleotides, transcription factors and other polypeptide transcriptional regulators, and non-macromolecular chemical entities which are capable of interacting with specific polynucleotide sequences. Effective compounds may alter polynucleotide expression by acting as either inhibitors or promoters of polynucleotide expression. Thus, in the treatment of disorders associated with increased PKIN
expression or activity, a compound which specifically inhibits expression of the polynucleotide encoding PKIN may be therapeutically useful, and in the treatment of disorders associated with decreased PKIN expression or activity, a compound which specifically promotes expression of the polynucleotide encoding PKIN may be therapeutically useful.
At least one, and up to a plurality, of test compounds may be screened for effectiveness in altering expression of a specific polynucleotide. A test compound may be obtained by any method commonly known in the art, including chemical modification of a compound known to be effective in altering polynucleotide expression; selection from an existing, commercially-available or proprietary library of naturally-occurnng or non-natural chemical compounds; rational design of a compound based on chemical and/or structural properties of the target polynucleotide;
and selection from a library of chemical compounds created combinatorially or randomly. A sample comprising a polynucleotide encoding PKIN is exposed to at least one test compound thus obtained. The sample may comprise, for example, an intact or permeabilized cell, or an in vitro cell-free or reconstituted biochemical system. Alterations in the expression of a polynucleotide encoding PI~IN are assayed by any method commonly known in the art. Typically, the expression of a specific nucleotide is detected by hybridization with a probe having a nucleotide sequence complementary to the sequence of the polynucleotide encoding PKIN. The amount of hybridization may be quantified, thus forming the basis for a comparison of the expression of the polynucleotide both with and without exposure to one or more test compounds. Detection of a change in the expression of a polynucleotide exposed to a test compound indicates that the test compound is effective in altering the expression of the polynucleotide. A screen for a.compound effective in altering expression of a specific polynucleotide can be carried out, for example, using a Schizosaccharomyces pombe gene expression system (Atkins, D. et al. (1999) U.S. Patent No. 5,932,435; Arndt, G.M. et al. (2000) Nucleic Acids Res. 28:E15) or a human cell fine such as HeLa cell (Clarke, M.L. et al. (2000) Biochem.
Biophys. Res. Commun.
268:8-13). A particular embodiment of the present invention involves screening a combinatorial library of oligonucleotides (such as deoxyribonucleotides, ribonucleotides, peptide nucleic acids, and modified oligonucleotides) for antisense activity against a specific polynucleotide sequence (Bruice, T.W. et al.
(1997) U.S. Patent No. 5,686,242; Bruice, T.W. et al. (2000) U.S. Patent No.
6,022,691).
Many methods for introducing vectors into cells or tissues are available and equally suitable for use in vivo, in vitro, and ex vivo. For ex vivo therapy, vectors may be introduced into stem cells taken from the patient and clonally propagated for autologous transplant back into that same patient.
Delivery by transfection, by liposome injections, or by polycationic amino polymers may be achieved using methods which are well known in the art. (See, e.g., Goldman, C.K. et al. (1997) Nat.
Biotechnol. 15:462-466.) Any of the therapeutic methods described above may be applied to any subject in need of such therapy, including, for example, mammals such as humans, dogs, cats, cows, horses, rabbits, and monkeys.
An additional embodiment of the invention relates to the administration of a composition which generally comprises an active ingredient formulated with a pharmaceutically acceptable excipient.
Excipients may include, for example, sugars, starches, celluloses, gums, and proteins. Various formulations are commonly known and are thoroughly discussed in the latest edition of Remin~ton's Pharmaceutical Sciences (Maack Publishing, Easton PA). Such compositions may consist of PKIN, antibodies to PKIN, and mimetics, agonists, antagonists, or inhibitors of P1~IN.
The compositions utilized in this invention may be administered by any number of routes including, but not limited to, oral, intravenous, intramuscular, infra-arterial, intramedullary, intrathecal, intraventricular, pulmonary, transdermal, subcutaneous, intraperitoneal, intranasal, enteral, topical, sublingual, or rectal means.
Compositions for pulmonary administration may be prepared in liquid or dry powder form.
These compositions are generally aerosolized immediately prior to inhalation by the patient. In the case of small molecules (e.g. traditional low molecular weight organic drugs), aerosol delivery of fast-acting formulations is well-known in the art. In the case of macromolecules (e.g. larger peptides and proteins), recent developments in the field of pulmonary delivery via the alveolar region of the lung have enabled the practical delivery of drugs such as insulin to blood circulation (see, e.g., Patton, J.S.
et al., U.S. Patent No. 5,997,848). Pulmonary delivery has the advantage of administration without needle injection, and obviates the need for potentially toxic penetration enhancers.
Compositions suitable for use in the invention include compositions wherein the active ingredients are contained in an effective amount to achieve the intended purpose. The determination of an effective dose is well within the capability of those skilled in the art.
Specialized forms of compositions may be prepared for direct intracellular delivery of macromolecules comprising PKIN or fragments thereof. For example, liposome preparations containing a cell-impermeable macromolecule may promote cell fusion and intracellular delivery of the macromolecule. Alternatively, PKIN or a fragment thereof may be joined to a short cationic N-terminal portion from the HIV Tat-1 protein. Fusion proteins thus generated have been found to transduce into the cells of all tissues, including the brain, in a mouse model system (Schwarze, S.R. et al. (1999) Science 285:1569-1572).
For any compound, the therapeutically effective dose can be estimated initially either in cell culture assays, e.g., of neoplastic cells, or in animal models such as mice, rats, rabbits, dogs, monkeys, or pigs. An animal model may also be used to determine the appropriate concentration range and route of administration. Such information can then be used to determine useful doses and routes for administration in humans.
A therapeutically effective dose refers to that amount of active ingredient, for example PK1N
or fragments thereof, antibodies of PKIN, and agonists, antagonists or inhibitors of PKIN, which ameliorates the symptoms or condition. Therapeutic efficacy and toxicity may be determined by standard pharmaceutical procedures in cell cultures or with experimental animals, such as by calculating the EDso (the dose therapeutically effective in 50% of the population) or LDso (the dose lethal to 50% of the population) statistics. The dose ratio of toxic to therapeutic effects is the therapeutic index, which can be expressed as the LDso/EDso ratio. Compositions which exhibit large therapeutic indices are preferred. The data obtained from cell culture assays and animal studies are used to formulate a range of dosage for human use. The dosage contained in such compositions is preferably within a range of circulating concentrations that includes the EDso with little or no toxicity.
The dosage varies within this range depending upon the dosage form employed, the sensitivity of the patient, and the route of administration.
The exact dosage will be determined by the practitioner, in light of factors related to the subject requiring treatment. Dosage and administration are adjusted to provide sufficient levels of the active moiety or to maintain the desired effect. Factors which may be taken into account include the severity of the disease state, the general health of the subject, the age, weight, and gender of the subject, time and frequency of administration, drug combination(s), reaction sensitivities, and response to therapy. Long-acting compositions may be administered every 3 to 4 days, every week, or biweekly depending on the half life and clearance rate of the particular formulation.
Normal dosage amounts may vary from about 0.1 ,ug to 100,000 ~cg, up to a total dose of about 1 gram, depending upon the route of administration. Guidance as to particular dosages and methods of delivery is provided in the literature and generally available to practitioners in the art.
Those skilled in the art will employ different formulations for nucleotides than for proteins or their inhibitors. Similarly, delivery of polynucleotides or polypeptides will be specific to particular cells, conditions, locations, etc.
DIAGNOSTICS
In another embodiment, antibodies which specifically bind PKIN may be used for the diagnosis of disorders characterized by expression of PKIN, or in assays to monitor patients being treated with PKIN or agonists, antagonists, or inhibitors of PKIN. Antibodies useful for diagnostic purposes may be prepared in the same manner as described above for therapeutics. Diagnostic assays for PKIN include methods which utilize the antibody and a label to detect PKIN in human body fluids or in extracts of cells or tissues. The antibodies, may be used with or without modification, and may be labeled by covalent or non-covalent attachment of a reporter molecule.
A wide variety of reporter molecules, several of which are described above, are known in the art and may be used.
A variety of protocols for measuring PKIN, including ELISAs, RIAs, and FACS, are known in the art and provide a basis for diagnosing altered or abnormal levels of PKIN expression. Normal or standard values for PKIN expression are established by combining body fluids or cell extracts taken from normal mammalian subjects, for example, human subjects, with antibodies to PKIN under conditions suitable for complex formation. The amount of standard complex formation may be quantitated by various methods, such as photometric means. Quantities of PKIN
expressed in subject, control, and disease samples from biopsied tissues are compared with the standard values. Deviation between standard and subject values establishes the parameters for diagnosing disease.
In another embodiment of the invention, the polynucleotides encoding PK1N may be used for diagnostic purposes. The polynucleotides which may be used include oligonucleotide sequences, complementary RNA and DNA molecules, and PNAs. The polynucleotides may be used to detect and quantify gene,expression in biopsied tissues in which expression of PKIN
may be correlated with disease. The diagnostic assay may be used to determine absence, presence, and excess expression of PK1N, and to monitor regulation of PKIN levels during therapeutic intervention.
In one aspect, hybridization with PCR probes which are capable of detecting polynucleotide sequences, including genomic sequences, encoding PK1N or closely related molecules may be used to identify nucleic acid sequences which encode PKIN. The specificity of the probe, whether it is made from a highly specific region, e.g., the 5'regulatory region, or from a less specific region, e.g., a conserved motif, and the stringency of the hybridization or amplification will determine whether the probe identifies only naturally occurring sequences encoding PKIN, allelic variants, or related sequences.
Probes may also be used for the detection of related sequences, and may have at least 50%
sequence identity to any of the PKIN encoding sequences. The hybridization probes of the subject invention may be DNA or RNA and may be derived from the sequence of SEQ ID
N0:21-40 or from genomic sequences including promoters, enhancers, and introns of the PK1N
gene.
Means for producing specific hybridization probes for DNAs encoding PK1N
include the cloning of polynucleotide sequences encoding PK1N or PKIN derivatives into vectors for the production of mRNA probes. Such vectors are known in the art, are commercially available, and may be used to synthesize RNA probes in vitro by means of the addition of the appropriate RNA
polymerases and the appropriate labeled nucleotides. Hybridization probes may be labeled by a variety of reporter groups, for example, by radionuclides such as 32P or 35S, or by enzymatic labels, such as alkaline phosphatase coupled to the probe via avidin/biotin coupling systems, and the like.
Polynucleotide sequences encoding PKIN may be used for the diagnosis of disorders associated with expression of PKIN. Examples of such disorders include, but are not limited to, a cancer, such as adenocarcinoma, leukemia, lymphoma, melanoma, myeloma, sarcoma, teratocarcinoma, and, in particular, cancers of the adrenal gland, bladder, bone, bone marrow, brain, breast, cervix, gall bladder, ganglia, gastrointestinal tract, heart, kidney, liver, lung, muscle, ovary, pancreas, parathyroid, penis, prostate, salivary glands, skin, spleen, testis, thymus, thyroid, and uterus, leukemias such as multiple myeloma and lymphomas such as Hodgkin's disease; an immune disorder, such as acquired immunodeficiency syndrome (AIDS), Addison's disease, adult respiratory distress syndrome, allergies, ankylosing spondylitis, amyloidosis, anemia, asthma, atherosclerosis, autoimmune hemolytic anemia, autoimmune thyroiditis, autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED), bronchitis, cholecystitis, contact dermatitis, Crohn's disease, atopic dermatitis, dermatomyositis, diabetes mellitus, emphysema, episodic lymphopenia with lymphocytotoxins, erythroblastosis fetalis, erythema nodosum, atrophic gastritis, glomerulonephritis, Goodpasture's syndrome, gout, Graves' disease, Hashimoto's thyroiditis, hypereosinophilia, irritable bowel syndrome, multiple sclerosis, myasthenia gravis, myocardial or pericardial inflammation, osteoarthritis, osteoporosis, pancreatitis, polymyositis, psoriasis, Reiter's syndrome, rheumatoid arthritis, scleroderma, Sjogren's syndrome, systemic anaphylaxis, systemic lupus erythematosus, systemic sclerosis, thrombocytopenic purpura, ulcerative colitis, uveitis, Werner syndrome, complications of cancer, hemodialysis, and extracorporeal circulation, viral, bacterial, fungal, parasitic, protozoal, and hehninthic infections, and trauma; a growth and developmental disorder, such as actinic keratosis, arteriosclerosis, atherosclerosis, bursitis, cirrhosis, hepatitis, mixed connective tissue disease (MCTD), myelofibrosis, paroxysmal nocturnal hemoglobinuria, polycythemia very, psoriasis, primary thrombocythemia, and cancers including adenocarcinoma, leukemia, lymphoma, melanoma, myeloma, sarcoma, teratocarcinoma, and, in particular, cancers of the adrenal gland, bladder, bone, bone marrow, brain, breast, cervix, gall bladder, ganglia, gastrointestinal tract, heart, kidney, liver, lung, muscle, ovary, pancreas, parathyroid, penis, prostate, salivary glands, skin, spleen, testis, thymus, thyroid, and uterus, renal tubular acidosis, anemia, Cushing's syndrome, achondroplastic dwarfism, Duchenne and Becker muscular dystrophy, epilepsy, gonadal dysgenesis, WAGR syndrome (Wilins' tumor, aniridia, genitourinary abnormalities, and mental retardation), Smith-Magenis syndrome, myelodysplastic syndrome, hereditary mucoepithelial dysplasia, hereditary keratodermas, hereditary neuropathies such as Charcot-Marie-Tooth disease and neurofibromatosis, hypothyroidism, hydrocephalus, seizure disorders such as Syndenham's chorea and cerebral palsy, spiny bifida, anencephaly, craniorachischisis, congenital glaucoma, cataract, and sensorineural hearing loss; a cardiovascular disease, such as arteriovenous fistula, atherosclerosis, hypertension, vasculitis, Raynaud's disease, aneurysms, arterial dissections, varicose veins, thrombophlebitis and phlebothrombosis, vascular tumors, and complications of thrombolysis, balloon angioplasty, vascular replacement, and coronary artery bypass graft surgery, congestive heart failure, ischemic heart disease, angina pectoris, myocardial infarction, hypertensive heart disease, degenerative valvular heart disease, calcific aortic valve stenosis, congenitally bicuspid aortic valve, mural annular calcification, mitral valve prolapse, rheumatic fever and rheumatic heart disease, infective endocarditis, nonbacterial thrombotic endocarditis, endocarditis of systemic lupus erythematosus, carcinoid heart disease, cardiomyopathy, myocarditis, pericarditis, neoplastic heart disease, congenital heart disease, and complications of cardiac transplantation, congenital lung anomalies, atelectasis, pulmonary congestion and edema, pulmonary embolism, pulmonary hemorrhage, pulmonary infarction, pulmonary hypertension, vascular sclerosis, obstructive pulmonary disease, restrictive pulmonary disease, chronic obstructive pulmonary disease, emphysema, chronuc bronchitis, bronchial asthma, bronchiectasis, bacterial pneumonia, viral and mycoplasmal pneumonia, lung abscess, pulmonary tuberculosis, diffuse interstitial diseases, pneumoconioses, sarcoidosis, idiopathic pulmonary fibrosis, desquamative interstitial pneumonitis, hypersensitivity pneumonitis, pulmonary eosinophilia bronchiolitis obliterans-organizing pneumonia, diffuse pulmonary hemorrhage syndromes, Goodpasture's syndromes, idiopathic pulmonary hemosiderosis, pulmonary involvement in collagen-vascular disorders, pulmonary alveolar proteinosis, lung tumors, inflammatory and noninflammatory pleural effusions, pneumothorax, pleural tumors, drug-induced lung disease, radiation-induced lung disease, and complications of lung transplantation; and a lipid disorder such as fatty liver, cholestasis, primary biliary cirrhosis, carnitine deficiency, carnitine palinitoyltransferase deficiency, myoadenylate deaminase deficiency, hypertriglyceridemia, lipid storage disorders such Fabry's disease, Gaucher's disease, Niemann-Pick's disease, metachromatic leukodystrophy, adrenoleukodystrophy, GM2 gangliosidosis, and ceroid lipofuscinosis, abetalipoproteinemia, Tangier disease, hyperlipoproteinemia, diabetes mellitus, lipodystrophy, lipomatoses, acute panniculitis, disseminated fat necrosis, adiposis dolorosa, lipoid adrenal hyperplasia, minimal change disease, lipomas, atherosclerosis, hypercholesterolemia, hypercholesterolemia with hypertriglyceridemia, primary hypoalphalipoproteinemia, hypothyroidism, renal disease, liver disease, lecithin:cholesterol acyltransferase deficiency, cerebrotendinous xanthomatosis, sitosterolemia, hypocholesterolemia, Tay-Sachs disease, Sandhoff s disease, hyperlipidemia, hyperlipemia, lipid myopathies, and obesity. The polynucleotide sequences encoding PKIN may be used in Southern or northern analysis, dot blot, or other membrane-based technologies; in PCR
technologies; in dipstick, pin, and multiformat ELISA-like assays; and in microarrays utilizing fluids or tissues from patients to detect altered PKIN expression. Such qualitative or quantitative methods are well known in the art.
In a particular aspect, the nucleotide sequences encoding PKIN may be useful in assays that detect the presence of associated disorders, particularly those mentioned above. The nucleotide sequences encoding PKIN may be labeled by standard methods and added to a fluid or tissue sample from a patient under conditions suitable for the formation of hybridization complexes. After a suitable incubation period, the sample is washed and the signal is quantified and compared with a standard value. If the amount of signal in the patient sample is significantly altered in comparison to a control sample then the presence of altered levels of nucleotide sequences encoding PKIN in the sample indicates the presence of the associated disorder. Such assays may also be used to evaluate the efficacy of a particular therapeutic treatment regimen in animal studies, in clinical trials, or to monitor the treatment of an individual patient.
In order to provide a basis for the diagnosis of a disorder associated with expression of PI~IN, a normal or standard profile for expression is established. This may be accomplished by combining body fluids or cell extracts taken from normal subjects, either animal or human, with a sequence, or a fragment thereof, encoding PI~IN, under conditions suitable for hybridization or amplification.
Standard hybridization may be quantified by comparing the values obtained from normal subjects with values from an experiment in which a known amount of a substantially purified polynucleotide is used.
Standard values obtained in this manner may be compared with values obtained from samples from patients who are symptomatic for a disorder. Deviation from standard values is used to establish the presence of a disorder.
Once the presence of a disorder is established and a treatment protocol is initiated, hybridization assays may be repeated on a regular basis to determine if the level of expression in the patient begins to approximate that which is observed in the normal subject.
The results obtained from successive assays may be used to show the efficacy of treatment over a period ranging from several.
days to months.
With respect to cancer, the presence of an abnormal amount of transcript (either under- or overexpressed) in biopsied tissue from an individual may indicate a predisposition for the development of the disease, or may provide a means for detecting the disease prior to the appearance of actual clinical symptoms. A more definitive diagnosis of this type may allow health professionals to employ preventative measures or aggressive treatment earlier thereby preventing the development or further progression of the cancer.
Additional diagnostic uses for oligonucleotides designed from the sequences encoding PKIN
may involve the use of PCR. These oligomers may be chemically synthesized, generated enzymatically, or produced in vitro. Oligomers will preferably contain a fragment of a polynucleotide encoding PKIN, or a fragment of a polynucleotide complementary to the polynucleotide encoding PKIN, and will be employed under optimized conditions for identification of a specific gene or condition. Oligomers may also be employed under less stringent conditions for detection or quantification of closely related DNA or RNA sequences.
In a particular aspect, oligonucleotide primers derived from the polynucleotide sequences encoding PKIN may be used to detect single nucleotide polymorphisms (SNPs).
SNPs are substitutions, insertions and deletions that are a frequent cause of inherited or acquired genetic disease in humans. Methods of SNP detection include, but are not limited to, single-stranded conformation polymorphism (SSCP) and fluorescent SSCP (fSSCP) methods. In SSCP, oligonucleotide primers derived from the polynucleotide sequences encoding PKIN are used to amplify DNA using the polymerase chain reaction (PCR). The DNA may be derived, for example, from diseased or normal tissue, biopsy samples, bodily fluids, and the like. SNPs in the DNA cause differences in the secondary and tertiary structures of PCR products in single-stranded form, and these differences are detectable using gel electrophoresis in non-denaturing gels. In fSCCP, the oligonucleotide primers are fluorescently labeled, which allows detection of the amplimers in high-throughput equipment such as DNA sequencing machines. Additionally, sequence database analysis methods, termed in silico SNP
(isSNP), are capable of identifying polymorphisms by comparing the sequence of individual overlapping DNA fragments which assemble into a common consensus sequence.
These computer-based methods filter out sequence variations due to laboratory preparation of DNA and sequencing errors using statistical models and automated analyses of DNA sequence chromatograms. In the alternative, SNPs may be detected and characterized by mass spectrometry using, for example, the high throughput MASSARRAY system (Sequenom, Inc., San Diego CA).
Methods which may also be used to quantify the expression of PKIN include radiolabeling or biotinylating nucleotides, coamplification of a control nucleic acid, and interpolating results from standard curves. (See, e.g., Melby, P.C. et al. (1993) J. Immunol. Methods 159:235-244; Duplaa, C.
et al. (1993) Anal. Biochem. 212:229-236.) The speed of quantitation of multiple samples may be accelerated by running the assay in a high-throughput format where the oligomer or polynucleotide of interest is presented in various dilutions and a spectrophotometric or colorimetric response gives rapid quantitation.
In further embodiments, oligonucleotides or longer fragments derived from any of the polynucleotide sequences described herein may be used as elements on a microarray. The microarray can be used in transcript imaging techniques which monitor the relative expression levels of large numbers of genes simultaneously as described below. The microarray may also be used to identify genetic variants, mutations, and polymorphisms. This information may be used to determine gene function, to understand the genetic basis of a disorder, to diagnose a disorder, to monitor progression/regression of disease as a function of gene expression, and to develop and monitor the activities of therapeutic agents in the treatment of disease. In particular, this information may be used to develop a pharmacogenomic profile of a patient in order to select the most appropriate and effective treatment regimen for that patient. For example, therapeutic agents which are highly effective and display the fewest side effects may be selected for a patient based on his/her pharmacogenomic profile.
In another embodiment, PI~IN, fragments of PKIN, or antibodies specific for PI~IN may be used as elements on a microarray. The microarray may be used to monitor or measure protein-protein interactions, drug-target interactions, and gene expression profiles,' as described above.
A particular embodiment relates to the use of the polynucleotides of the present invention to generate a transcript image of a tissue or cell type. A transcript image represents the global pattern of gene expression by a particular tissue or cell type. Global gene expression patterns are analyzed by quantifying the number of expressed genes and their relative abundance under given conditions and at a given time. (See Seilhamer et al., "Comparative Gene Transcript Analysis,"
U.S. Patent Number 5,840,484, expressly incorporated by reference herein.) Thus a transcript image may be generated by hybridizing the polynucleotides of the present invention or their complements to the totality of transcripts or reverse transcripts of a particular tissue or cell type. In one embodiment, the hybridization takes place in high-throughput format, wherein the polynucleotides of the present invention or their complements comprise a subset of a plurality of elements on a microarray. The resultant transcript image would provide a profile~of gene activity.
Transcript images may be generated using transcripts isolated from tissues, cell lines, biopsies, or other biological samples. The transcript image may thus reflect gene expression in vivo, as in the case of a tissue or biopsy sample, or in vitro, as in the case of a cell fine.
Transcript images which profile the expression of the polynucleotides of the present invention may also be used in conjunction with in vitro model systems and preclinical evaluation of pharmaceuticals, as well as toxicological testing of industrial and naturally-occurring environmental compounds. All compounds induce characteristic gene expression patterns, frequently termed molecular fingerprints or toxicant signatures, which are indicative of mechanisms of action and toxicity (Nuwaysir, E.F. et al. (1999) Mol. Carcinog. 24:153-159; Steiner, S. and N.L.
Anderson (2000) Toxicol. Lett. 112-113:467-471, expressly incorporated by reference herein).
If a test compound has a signature similar to that of a compound with known toxicity, it is likely to share those toxic properties.
These fingezprints or signatures are most useful and refined when they contain expression information from a large number of genes and gene families. Ideally, a genome-wide measurement of expression provides the highest quality signature. Even genes whose expression is not altered by any tested compounds are important as well, as the levels of expression of these genes are used to normalize the rest of the expression data. The normalization procedure is useful for comparison of expression data after treatment with different compounds. While the assignment of gene function to elements of a toxicant signature aids in interpretation of toxicity mechanisms, knowledge of gene function is not necessary for the statistical matching of signatures which leads to prediction of toxicity. (See, for example, Press Release 00-02 from the National Institute of Environmental Health Sciences, released February 29, 2000, available at http:/lwww.niehs.nih.gov/oc/newsltoxchip.htm.) 'Therefore, it is important and desirable in toxicological screening using toxicant signatures to include all expressed gene sequences.
In one embodiment, the toxicity of a test compound is assessed by treating a biological sample containing nucleic acids with the test compound. Nucleic acids that are expressed in the treated biological sample are hybridized with one or more probes specific to the polynucleotides of the present invention, so that transcript levels corresponding to the polynucleotides of the present invention may be quantified. The transcript levels in the treated biological sample are compared with levels in an untreated biological sample. Differences in the transcript levels between the two samples are indicative of a toxic response caused by the test compound in the treated sample.
Another particular embodiment relates to the use of the polypeptide sequences of the present invention to analyze the proteome of a tissue or cell type. The term proteome refers to.the global pattern of protein expression in a particular tissue or cell type. Each protein component of a proteome can be subjected individually to further analysis. Proteome expression patterns, or profiles, are analyzed by quantifying the number of expressed proteins and their relative abundance under given conditions and at a given time. A profile of a cell's proteome may thus be generated by separating and analyzing the polypeptides of a particular tissue or cell type. In one embodiment, the separation is achieved using two-dimensional gel electrophoresis, in which proteins from a sample are separated by isoelectric focusing in the first dimension, and then according to molecular weight by sodium dodecyl sulfate slab gel electrophoresis in the second dimension (Steiner and Anderson, s-u~ra). The proteins are visualized in the gel as discrete and uniquely positioned spots, typically by staining the gel with an agent such as Coomassie Blue or silver or fluorescent stains. The optical density of each protein spot is generally proportional to the level of the protein in the sample. The optical densities of equivalently positioned protein spots from different samples, for example, from biological samples either treated or untreated with a test compound or therapeutic agent, are compared to identify any changes in protein spot density related to the treatment. The proteins in the spots are partially sequenced using, for example, standard methods employing chemical or enzymatic cleavage followed by mass spectrometry. The identity of the protein in a spot may be determined by comparing its partial sequence, preferably of at least 5 contiguous amino acid residues, to the polypeptide sequences of the present invention. In some cases, further sequence data may be obtained for definitive protein identification.

A proteomic profile may also be generated using antibodies specific for PKIN
to quantify the levels of PKIN expression. In one embodiment, the antibodies are used as elements on a microarray, and protein expression levels are quantified by exposing the microarray to the sample and detecting the levels of protein bound to each array element (Lueking, A. et al. (1999) Anal. Biochem. 270:103-111; Mendoze, L.G, et al. (1999) Biotechniques 27:778-788). Detection may be performed by a variety of methods known in the art, for example, by reacting the proteins in the sample with a thiol- or amino-reactive fluorescent compound and detecting the amount of fluorescence bound at each array element.
Toxicant signatures at the proteome level are also useful for toxicological screening, and l0 should be analyzed in parallel with toxicant signatures at the transcript level. There is a poor correlation between transcript and protein abundances for some proteins in some tissues (Anderson, N.L. and J. Seilhamer (1997) Electrophoresis 18:533-537), so proteome toxicant signatures may be useful in the analysis of compounds which do not significantly affect the transcript image, but which alter the proteomic profile. In addition, the analysis of transcripts in body fluids is difficult, due to rapid degradation of mRNA, so proteomic profiling may be more reliable and informative in such cases.
In another embodiment, the toxicity of a test compound is assessed by treating a biological sample containing proteins with the test compound. Proteins that are expressed in the treated biological sample are separated so that the amount of each protein can be quantified. The amount of each protein is compared to the amount of the corresponding protein in an untreated biological sample.
A difference in the amount of protein between the two samples is indicative of a toxic response to the test compound in the treated sample. Individual proteins are identified by sequencing the amino acid residues of the individual proteins and comparing these partial sequences to the polypeptides of the present invention.
In another embodiment, the toxicity of a test compound is assessed by treating a biological sample containing proteins with the test compound. Proteins from the biological sample are incubated with antibodies specific to the polypeptides of the present invention. The amount of protein recognized by the antibodies is quantified. The amount of protein in the treated biological sample is compared with the amount in an untreated biological sample. A difference in the amount of protein between the two samples is indicative of a toxic response to the test compound in the treated sample.
Microarrays may be prepared, used, and analyzed using methods known in the art. (See, e.g., Brennan, T.M. et al. (1995) U.S. Patent No. 5,474,796; Schena, M. et al.
(1996) Proc. Natl. Acad.
Sci. USA 93:10614-10619; Baldeschweiler et al. (1995) PCT application W095/251116; Shalon, D. et al. (1995) PCT application W095/35505; Heller, R.A, et al. (1997) Proc. Natl.
Acad. Sci. USA

94:2150-2155; and Heller, M.J. et al. (1997) U.S. Patent No. 5,605,662.) Various types of microarrays are well known and thoroughly described in DNA Microarrays: A
Practical Approach, M. Schena, ed. (1999) Oxford University Press, London, hereby expressly incorporated by reference.
In another embodiment of the invention, nucleic acid sequences encoding PKIN
may be used to generate hybridization probes useful in mapping the naturally occurring genomic sequence. Either coding or noncoding sequences may be used, and in some instances, noncoding sequences may be preferable over coding sequences. For example, conservation of a coding sequence among members of a multi-gene family may potentially cause undesired cross hybridization during chromosomal mapping. The sequences may be mapped to a particular chromosome, to a specific region of a chromosome, or to artificial chromosome constructions, e.g., human artificial chromosomes (HACs), yeast artificial chromosomes (YACs), bacterial artificial chromosomes (BACs), bacterial P1 constructions, or single chromosome cDNA libraries. (See, e.g., Harnngton, J.J. et al. (1997) Nat.
Genet. 15:345-355; Price, C.M. (1993) Blood Rev. 7:127-134; and Trask, B.J.
(1991) Trends Genet.

7:149-154.) Once mapped, the nucleic acid sequences of the invention may be used to develop genetic linkage maps, for example, which correlate the inheritance of a disease state with the inheritance of a particular chromosome region or restriction fragment length polymorphism (RFLP):
(See, for example, Lander, E.S. and D. Botstein (1986) Proc. Natl. Acad. Sci.
USA 83:7353-7357.) Fluorescent in situ hybridization (FISH) may be correlated with other physical and genetic map data. (See, e.g., Heinz-Ulrich, et al. (1995) in Meyers, supra, pp. 965-968.) Examples of genetic map data can be found in various scientific journals or at the Online Mendelian Inheritance in Man (OMIM) World Wide Web site. Correlation between the location of the gene encoding PKIN on a physical map and a specific disorder, or a predisposition to a specific disorder, may help define the region of DNA associated with that disorder and thus may further positional cloning efforts.
In situ hybridization of chromosomal preparations and physical mapping techniques, such as linkage analysis using established chromosomal markers, may be used for extending genetic maps.
Often the placement of a gene on the chromosome of another mammalian species, such as mouse, may reveal associated markers even if the exact chromosomal locus is not known. This information is valuable to investigators searching for disease genes using positional cloning or other gene discovery techniques. Once the gene or genes responsible for a disease or syndrome have been crudely localized by genetic linkage to a particular genomic region, e.g., ataxia-telangiectasia to 11q22-23, any sequences mapping to that area may represent associated or regulatory genes for further investigation.
(See, e.g., Gatti, R.A. et al. (1988) Nature 336:577-580.) The nucleotide sequence of the instant invention may also be used to detect differences in the chromosomal location due to translocation, inversion, etc., among normal, carrier, or affected individuals.
In another embodiment of the invention, PKIN, its catalytic or immunogenic fragments, or oligopeptides thereof can be used for screening libraries of compounds in any of a variety of drug screening techniques. The fragment employed in such screening may be free in solution, affixed to a solid support, borne on a cell surface, or located intracellularly. The formation of binding complexes between PKIN and the agent being tested may be measured.
Another technique for drug screening provides for high throughput screening of compounds having suitable binding affinity to the protein of interest. (See, e.g., Geysen, et al. (1984) PCT
application W084/03564.) In this method, Large numbers of different small test compounds are synthesized on a solid substrate. The test compounds are reacted with PI~IN, or fragments thereof, and washed. Bound PKIN is then detected by methods well known in the art.
Purified PI~IN can also be coated directly onto plates for use in the aforementioned drug screening techniques.
Alternatively, non-neutralizing antibodies can be used to capture the peptide and immobilize it on a solid support.
In another embodiment, one may use competitive drug screening assays in which neutralizing antibodies capable of binding PKIN specifically compete with a test compound for binding PKIN. In this manner, antibodies can be used to detect the presence of any peptide which shares one or more antigenic determinants with PKIN.
In additional embodiments, the nucleotide sequences which encode PKIN may be used in any molecular biology techniques that have yet to be developed, provided the new techniques rely on properties of nucleotide sequences that are currently known, including, but not limited to, such properties as the triplet genetic code and specific base pair interactions.
Without further elaboration, it is believed that one skilled in the art can, using the preceding description, utilize the present invention to its fullest extent. The following embodiments are, therefore, to be construed as merely illustrative, and not limitative of the remainder of the disclosure in any way whatsoever.
The disclosures of all patents, applications, and publications mentioned above and below, including U.S. Ser. No. 60/220,038, U.S. Ser. No. 60/222,112, U.S. Ser. No.
60/222,831, and U.S. Ser.
No. 60/224,729 are hereby expressly incorporated by reference.
EXAMPLES
I. Construction of cDNA Libraries Incyte cDNAs were derived from cDNA libraries described in the LIFESEQ GOLD

database (Incyte Genomics, Palo Alto CA) and shown in Table 4, column 5. Some tissues were homogenized and lysed in guanidinium isothiocyanate, while others were homogenized and lysed in phenol or in a suitable mixture of denaturants, such as TRIZOL (Life Technologies), a monophasic solution of phenol and guanidine isothiocyanate. The resulting lysates were centrifuged over CsCl cushions or extracted with chloroform. RNA was precipitated from the lysates with either isopropanol or sodium acetate and ethanol, or by other routine methods.
Phenol extraction and precipitation of RNA were repeated as necessary to increase RNA
purity. In some cases, RNA was treated with DNase. For most libraries, poly(A)+ RNA was isolated using oligo d(T)-coupled paramagnetic particles (Promega), OLIGOTEX
latex particles (QIAGEN, Chatsworth CA), or an OLIGOTEX mRNA purification kit (QIAGEN).
Alternatively, RNA was isolated directly from tissue lysates using other RNA isolation kits, e.g., the POLY(A)PURE mRNA purification kit (Ambion, Austin TX).
In some cases, Stratagene was provided with RNA and constructed the corresponding cDNA
libraries. Otherwise, cDNA was synthesized and cDNA libraries were constructed with the UNIZAP vector system (Stratagene) or SUPERSCRIPT plasmid system (Life Technologies), using the recommended procedures or similar methods known in the art. (See, e.g., Ausubel, 1997, supra, units 5.1-6.6.) Reverse transcription was initiated using oligo d(T) or random primers. Synthetic oligonucleotide adapters were ligated to double stranded cDNA, and the cDNA
was digested with the appropriate restriction enzyme or enzymes. For most libraries, the cDNA was size-selected (300-1000 bp) using SEPHACRYL S 1000, SEPHAROSE CL2B, or SEPHAROSE CLA.B column chromatography (Amersham Pharmacia Biotech) or preparative agarose gel electrophoresis. cDNAs were ligated into compatible restriction enzyme sites of the polylinker of a suitable plasmid, e.g., PBLUESCRIPT plasmid (Stratagene), PSPORTl plasmid (Life Technologies), PCDNA2.1 plasmid (Invitrogen, Carlsbad CA), PBK-CMV plasmid (Stratagene), or pINCY (Incyte Genomics, Palo Alto CA), or derivatives thereof. Recombinant plasmids were transformed into competent E. coli cells including XL1-Blue, XLl-BIueMRF, or SOLR from Stratagene or DHSa, DH10B, or ElectroMAX
DH10B from Life Technologies.
II. Isolation of cDNA Clones Plasmids obtained as described in Example I were recovered from host cells by in vivo excision using the UNIZAP vector system (Stratagene) or by cell lysis.
Plasmids were purified using at least one of the following: a Magic or WIZARD Minipreps DNA purification system (Promega); an AGTC Miniprep purification kit (Edge Biosystems, Gaithersburg MD); and QIAWELL

8 Plasmid, QIAWELL 8 Plus Plasmid, QIAWELL 8 Ultra Plasmid purification systems or the R.E.A.L. PREP

96 plasmid purification kit from QIAGEN. Following precipitation, plasmids were resuspended in 0.1 ml of distilled water and stored, with or without lyophilization, at 4°C.
Alternatively, plasmid DNA was amplified from host cell lysates using direct link PCR in a high-throughput format (Rao, V.B. (1994) Anal. Biochem. 216:1-14). Host cell lysis and thermal cycling steps were carried out in a single reaction mixture. Samples were processed and stored in 384-well plates, and the concentration of amplified plasmid DNA was quantified fluorometrically using PICOGREEN dye (Molecular Probes, Eugene OR) and a FLUOROSKAN II fluorescence scanner (Labsystems Oy, Helsinki, Finland).
III. Sequencing and Analysis Incyte cDNA recovered in plasmids as described in Example II were sequenced as follows.
Sequencing reactions were processed using standard methods or high-throughput instrumentation such as the ABI CATALYST 800 (Applied Biosystems) thermal cycler or the PTC-200 thermal cycler (MJ Research) in conjunction with the HYDRA microdispenser (Robbins Scientific) or the MICROLAB 2200 (Hamilton) liquid transfer system. cDNA sequencing reactions were prepared using reagents provided by Amersham Pharmacia Biotech or supplied in ABI
sequencing kits such as the ABI PRISM BIGDYE Terminator cycle sequencing ready reaction kit (Applied Biosystems).
Electrophoretic separation of cDNA sequencing reactions and detection of labeled polynucleotides were carried out using the MEGABACE 1000 DNA sequencing system (Molecular Dynamics); the ABI PRISM 373 or 377 sequencing system (Applied Biosystems) in conjunction with standard ABI
protocols and base calling software; or other sequence analysis systems known in the art. Reading frames within the cDNA sequences were identified using standard methods (reviewed in Ausubel, 1997, supra, unit 7.7). Some of the cDNA sequences were selected for extension using the . techniques disclosed in Example VIII.
The polynucleotide sequences derived from Incyte cDNAs were validated by removing vector, linker, and poly(A) sequences and by masking ambiguous bases, using algorithms and programs based on BLAST, dynamic programming, and dinucleotide nearest neighbor analysis. The Incyte cDNA sequences or translations thereof were then queried against a selection of public databases such as the GenBank primate, rodent, mammalian, vertebrate, and eukaryote databases, and BLOCKS, PRINTS, DOMO, PRODOM, and hidden Markov model (HMM)-based protein family databases such as PFAM. (HMM is a probabilistic approach which analyzes consensus primary structures of gene families. See, for example, Eddy, S.R. (1996) Curr. Opin.
Struct. Biol. 6:361-365.) The queries were performed using programs based on BLAST, FASTA, BLIMPS, and HMMER.
The Incyte cDNA sequences were assembled to produce full length polynucleotide sequences.

Alternatively, GenBank cDNAs, GenBank ESTs, stitched sequences, stretched sequences, or Genscan-predicted coding sequences (see Examples IV and V~ were used to extend Incyte cDNA
assemblages to full length. Assembly was performed using programs based on Phred, Phrap, and Consed, and cDNA assemblages were screened for open reading frames using programs based on GeneMark, BLAST, and FASTA. The full length polynucleotide sequences were translated to derive the corresponding full length polypeptide sequences. Alternatively, a polypeptide of the invention may begin at any of the methionine residues of the full length translated polypeptide. Full length polypeptide sequences were subsequently analyzed by querying against databases such as the GenBankprotein databases (genpept), SwissProt, BLOCKS, PRINTS, DOMO, PRODOM, Prosite, and hidden Markov model (HMM)-based protein family databases such as PFAM. Full length polynucleotide sequences are also analyzed using MACDNASIS PRO software (Hitachi Software Engineering, South San Francisco CA) and LASERGENE software (DNASTAR). Polynucleotide and polypeptide sequence alignments are generated using default parameters specified by the CLUSTAL algorithm as incorporated into the MEGALIGN multisequence alignment program (DNASTAR), which also calculates the percent identity between aligned sequences.
Table 7 summarizes the tools, programs, and algorithms used for the analysis and assembly of Incyte cDNA and full length sequences and provides applicable descriptions, references, and threshold parameters. The first column of Table 7 shows the tools, programs, and algorithms used, the second column provides brief descriptions thereof, the third column presents appropriate references, all of which are incorporated by reference herein in their entirety, and the fourth column presents, where applicable, the scores, probability values, and other parameters used to evaluate the strength of a match between two sequences (the higher the score or the lower the probability value, the greater the identity between two sequences).
The programs described above for the assembly and analysis of full length polynucleotide and polypeptide sequences were also used to identify polynucleotide sequence fragments from SEQ ID
N0:21-40. Fragments from about 20 to about 4000 nucleotides which axe useful in hybridization and amplification technologies are described in Table 4, column 4.
IV. Identification and Editing of Coding Sequences from Genomic DNA
Putative human kinases were initially identified by running the Genscan gene identification .
program against public genomic sequence databases (e.g., gbpri and gbhtg).
Genscan is a general-purpose gene identification program which analyzes genomic DNA sequences from a variety of organisms (See Burge, C. and S. Karlin (1997) J. Mol. Biol. 268:78-94, and Burge, C. and S. Karlin (1998) Curr. Opin. Struct. Biol. 8:346-354). The program concatenates predicted exons to form an assembled cDNA sequence extending from a methionine to a stop codon. The output of Genscan is a FASTA database of polynucleotide and polypeptide sequences. The maximum range of sequence for Genscan to analyze at once was set to 30 kb. To determine which of these Genscan predicted cDNA
sequences encode human kinases, the encoded polypeptides were analyzed by querying against PFAM models for human kinases. Potential human kinases were also identified by homology to Incyte cDNA sequences that had been annotated as human kinases. These selected Genscan-predicted sequences were then compared by BLAST analysis to the genpept and gbpri public databases. Where necessary, the Genscan-predicted sequences were then edited by comparison to the top BLAST hit from genpept to correct errors in the sequence predicted by Genscan, such as l0 extra or omitted exons. BLAST analysis was also used to find any Incyte cDNA or public cDNA
coverage of the Genscan-predicted sequences, thus providing evidence for transcription. When Incyte cDNA coverage was available, this information was used to correct or confirm the Genscan predicted sequence. Full length polynucleotide sequences were obtained by assembling Genscan-predicted coding sequences with Incyte cDNA sequences and/or public cDNA sequences using the assembly process described in Example III. Alternatively, full length polynucleotide sequences were derived entirely from edited or unedited Genscan-predicted coding sequences.
V. Assembly of Genomic Sequence Data with cDNA Sequence Data "Stitched" Sequences Partial cDNA sequences were extended with exons predicted by the Genscan gene identification program described in Example IV. Partial cDNAs assembled as described in Example III were mapped to genomic DNA and parsed into clusters containing related cDNAs and Genscan exon predictions from one or more genomic sequences. Each cluster was analyzed using an algorithm based on graph theory and dynamic programming to integrate cDNA and genomic information, generating possible splice variants that were subsequently confirmed, edited, or extended to create a full length sequence. Sequence intervals in which the entire length of the interval was present on more than one sequence in the cluster were identified, and intervals thus identified were considered to be equivalent by transitivity. For example, if an interval was present on a cDNA and two genomic sequences, then all three intervals were considered to be equivalent. This process allows unrelated but consecutive genomic sequences to be brought together, bridged by cDNA
sequence. Intervals thus identified were then "stitched" together by the stitching algorithm in the order that they appear along their parent sequences to generate the longest possible sequence, as well as sequence variants.
Linkages between intervals which proceed along one type of parent sequence (cDNA to cDNA or genomic sequence to genomic sequence) were given preference over linkages which change parent type (cDNA to genomic sequence). The resultant stitched sequences were translated and compared by BLAST analysis to the genpept and gbpri public databases. Incorrect exons predicted by Genscan were corrected by comparison to the top BLAST hit from genpept. Sequences were further extended with additional cDNA sequences, or by inspection of genomic DNA, when necessary.
"Stretched" Sequences Partial DNA sequences were extended to full length with an algorithm based on BLAST
analysis. First, partial cDNAs assembled as described in Example III were queried against public databases such as the GenBank primate, rodent, mammalian, vertebrate, and eukaryote databases using the BLAST program. The nearest GenBank protein homolog was then compared by BLAST
analysis to either Incyte cDNA sequences or GenScan exon predicted sequences described in Example IV. A chimeric protein was generated by using the resultant high-scoring segment pairs (HSPs) to map the translated sequences onto the GenBank protein homolog.
Insertions or deletions may occur in the chimeric protein with respect to the original GenB ank protein homolog. The GenBank protein homolog, the chimeric protein, or both were used as probes to search for homologous genomic sequences from the public human genome databases. Partial DNA
sequences were therefore "stretched" ox extended by the addition of homologous genomic sequences. The resultant stretched sequences were examined to determine whether it contained a complete gene.
VI. Chromosomal Mapping of PKIN Encoding Polynucleotides The sequences which were used to assemble SEQ ID NO:21-40 were compared with sequences from the Incyte LIFESEQ database and public domain databases using BLAST and other implementations of the Smith-Waterman algorithm. Sequences from these databases that matched SEQ ID N0:21-40 were assembled into clusters of contiguous and overlapping sequences using assembly algorithms such as Phrap (Table 7). Radiation hybrid and genetic mapping data available from public resources such as the Stanford Human Genome Center (SHGC), Whitehead Institute for Genome Research (WIGR), and Genethon were used to determine if any of the clustered sequences had been previously mapped. Inclusion of a mapped sequence in a cluster resulted ir1 the assignment of all sequences of that cluster, including its particular SEQ ID NO:, to that map location.
Map locations are represented by ranges, or intervals, of human chromosomes.
The map position of an interval, in centiMorgans, is measured relative to the terminus of the chromosome's p-arm. (The centiMorgan (cM) is a unit of measurement based on recombination frequencies between chromosomal markers. On average, 1 cM is roughly equivalent to 1 megabase (Mb) of DNA in humans, although this can vary widely due to hot and cold spots of recombination.) The cM distances are based on genetic markers mapped by Genethon which provide boundaries for radiation hybrid markers whose sequences were included in each of the clusters. Human genome maps and other resources available to the public, such as the NCBI "GeneMap'99" World Wide Web site (http://www.ncbi.nlm.nih.gov/genemapn, can be employed to determine if previously identified disease genes map within or in proximity to the intervals indicated above.
VII. Analysis of Polynucleotide Expression Northern analysis is a laboratory technique used to detect the presence of a transcript of a gene and involves the hybridization of a labeled nucleotide sequence to a membrane on which RNAs from a particular cell type or tissue have been bound. (See, e.g., Sambrook, supra, ch. 7; Ausubel (1995) supra, ch. 4 and 16.) Analogous computer techniques applying BLAST were used to search for identical or related molecules in cDNA databases such as GenBank or LIFESEQ (Incyte Genomics). This analysis is much faster than multiple membrane-based hybridizations. In addition, the sensitivity of the computer search can be modified to determine whether any particular match is categorized as exact or similar.
The basis of the search is the product score, which is defined as:
BLAST Score x Percent Identity 5 x minimum { length(Seq. 1), length(Seq. 2) }
The product score takes into account both the degree of similarity between two sequences and the length of the sequence match. The product score is a normalized value between 0 and 100, and is calculated as follows: the BLAST score is multiplied by the percent nucleotide identity and the product is divided by (5 times the length of the shorter of the two sequences). The BLAST score is calculated by assigning a score of +5 for every base that matches in a high-scoring segment pair (HSP), and -4 for every mismatch. Two sequences may share more than one HSP
(separated by gaps). If there is more than one HSP, then the pair with the highest BLAST
score is used to calculate the product score. The product score represents a balance between fractional overlap and quality in a BLAST alignment. For example, a product score of 100 is produced only for 100%
identity over the entire length of the shorter of the two sequences being compared. A product score of 70 is produced, either by 100% identity and 70% overlap at one end, or by 88% identity and 100% overlap at the other. A product score of 50 is produced either by 100% identity and 50%
overlap at one end, or 79%
identity and 100% overlap.
Alternatively, polynucleotide sequences encoding PKIN are analyzed with respect to the tissue sources from which they were derived. For example, some full length sequences are assembled, at least in part, with overlapping Incyte cDNA sequences (see Example III). Each cDNA
sequence is derived from a cDNA library constructed from a human tissue. Each human tissue is classified into one of the following organ/tissue categories: cardiovascular system; connective tissue;
digestive system; embryonic structures; endocrine system; exocrine glands;
genitalia, female; genitalia, male; germ cells; heroic and immune system; liver; musculoskeletal system;
nervous system;
pancreas; respiratory system; sense organs; skin; stomatognathic system;
unclassified/mixed; or urinary tract. The number of libraries in each category is counted and divided by the total number of libraries across all categories. Similarly, each human tissue is classified into one of the following disease/condition categories: cancer, cell line, developmental, inflammation, neurological, trauma, cardiovascular, pooled, and other, and the number of libraries in each category is counted and divided by the total number of libraries across all categories. The resulting percentages reflect the tissue- and disease-specific expression of cDNA encoding PI~IN. cDNA sequences and cDNA
libraryltissue information are found in the LIFESEQ GOLD database (Incyte Genomics, Palo Alto CA).
VIII. Extension of PKIN Encoding Polynucleotides Full length polynucleotide sequences were also produced by extension of an appropriate fragment of the full length molecule using oligonucleotide primers designed from this fragment. One primer was synthesized to initiate 5' extension of the known fragment, and the other primer was synthesized to initiate 3' extension of the known fragment. The initial primers were designed using OLIGO 4.06 software (National Biosciences), or another appropriate program, to be about 22 to 30 nucleotides in length, to have a GC content of about 50% or more, and to anneal to the target sequence at temperatures of about 68°C to about 72°C. Any stretch of nucleotides which would result in hairpin structures and primer-primer dimerizations was avoided.
Selected human cDNA libraries were used to extend the sequence. If more than one extension was necessary or desired, additional or nested sets of primers were designed.
High fidelity amplification was obtained by PCR using methods well known in the art. PCR
was performed in 96-well plates using the PTC-200 thermal cycler (MJ Research, Inc.). The reaction mix contained DNA template, 200 nmol of each primer, reaction buffer containing Mg2+, (NH4)2S04, and 2-mercaptoethanol, Taq DNA polymerase (Amersham Pharmacia Biotech), ELONGASE
enzyme (Life Technologies), and Pfu DNA polymerase (Stratagene), with the following parameters for primer pair PCI A and PCI B: Step 1: 94°C, 3 min; Step 2:
94°C, 15 sec; Step 3: 60°C, 1 min;
Step 4: 68 °C, 2 min; Step 5: Steps 2, 3, and 4 repeated 20 times; Step 6: 68 °C, 5 min; Step 7: storage at 4°C. In the alternative, the parameters for primer pair T7 and SI~+
were as follows: Step 1: 94°C, 3 min; Step 2: 94°C, 15 sec; Step 3: 57°C, 1 min; Step 4:
68°C, 2 min; Step 5: Steps 2, 3, and 4 repeated 20 times; Step 6: 68 ° C, 5 min; Step 7: storage at 4 °
C.
The concentration of DNA in each well was determined by dispensing 100 ~1 PICOGREEN
quantitation reagent (0.25% (v/v) PICOGREEN; Molecular Probes, Eugene OR) dissolved in 1X TE
and 0.5 ~1 of undiluted PCR product into each well of an opaque fluorimeter plate (Corning Costar, Acton MA), allowing the DNA to bind to the reagent. The plate was scanned in a Fluoroskan II
(Labsystems Oy, Helsinki, Finland) to measure the fluorescence of the sample and to quantify the concentration of DNA. A 5 ~cl to 10 ,ul aliquot of the reaction mixture was analyzed by electrophoresis on a 1 % agarose gel to determine which reactions were successful in extending the sequence.
The extended nucleotides were desalted and concentrated, transferred to 384-well plates, digested with CviJI cholera virus endonuclease (Molecular Biology Research, Madison W17, and sonicated or sheared prior to religation into pUC 18 vector (Amersham Pharmacia Biotech). For shotgun sequencing, the digested nucleotides were separated on low concentration (0.6 to 0.8%) agarose gels, fragments were excised, and agar digested with Agar ACE
(Promega). Extended clones were religated using T4 lipase (New England Biolabs, Beverly MA) into pUC 18 vector (Amersham Pharmacia Biotech), treated with Pfu DNA polymerase (Stratagene) to fill=in restriction site overhangs, and transfected into competent E. coli cells. Transformed cells were selected on antibiotic-containing media, and individual colonies were picked and cultured overnight at 37 °C in 384-well plates in LB/2x carb liquid media.
The cells were lysed, and DNA was amplified by PCR using Taq DNA polymerase (Amersham Pharmacia Biotech) and Pfu DNA polymerase (Stratagene) with the following parameters: Step 1: 94 ° C, 3 min; Step 2: 94 ° C, 15 sec; Step 3: 60 ° C, 1 min; Step 4: 72 ° C, 2 min; Step 5: steps 2, 3, and 4 repeated 29 times; Step 6: 72°C, 5 min; Step 7:
storage at 4°C. DNA was quantified by PICOGREEN reagent (Molecular Probes) as described above. Samples with low DNA
recoveries were reamplified using the same conditions as described above.
Samples were diluted with 20% dimethysulfoxide (1:2, v/v), and sequenced using DYENAMIC energy transfer sequencing primers and the DYENAMIC DIRECT kit (Amersham Pharmacia Biotech) or the ABI
PRISM
BIGDYE Terminator cycle sequencing ready reaction kit (Applied Biosystems).
In like manner, full length polynucleotide sequences are verified using the above procedure or are used to obtain 5' regulatory sequences using the above procedure along with oligonucleotides designed for such extension, and an appropriate genomic library.
IX. Labeling and Use of Individual Hybridization Probes Hybridization probes derived from SEQ m N0:21-40 are employed to screen cDNAs, genomic DNAs, or mRNAs. Although the labeling of oligonucleotides, consisting of about 20 base pairs, is specifically described, essentially the same procedure is used with larger nucleotide fragments. Oligonucleotides are designed using state-of the-art software such as OLIGO 4.06 software (National Biosciences) and labeled by combining 50 pmol of each oligomer, 250 ~Ci of [y-32P] adenosine triphosphate (Amersham Pharmacia Biotech), and T4 polynucleotide kinase (DuPont NEN, Boston MA). The labeled oligonucleotides are substantially purified using a SEPHADEX G-25 supe~ne size exclusion dextran bead column (Amersham Pharmacia Biotech).
An aliquot containing 10' counts per minute of the labeled probe is used in a typical membrane-based hybridization analysis of human genomic DNA digested with one of the following endonucleases: Ase I, Bgl II, Eco RI, Pst I, Xba I, or Pvu II (DuPont NEN).
The DNA from each digest is fractionated on a 0.7% agarose gel and transferred to nylon membranes (Nytran Plus, Schleicher & Schuell, Durham NH). Hybridization is carried out for 16 hours at 40°C. To remove nonspecific signals, blots are sequentially washed at room temperature under conditions of up to, for example, 0.1 x saline sodium citrate and 0.5%
sodium dodecyl sulfate.
Hybridization patterns axe visualized using autoradiography or an alternative imaging means and compared.
X. Microarrays The linkage or synthesis of array elements upon a microarray can be achieved utilizing.
photolithography, piezoelectric printing (ink jet printing, See, e.g., Baldeschweiler, supra.), mechanical microspotting technologies, and derivatives thereof. The substrate in each of the aforementioned technologies should be uniform and solid with a non-porous surface (Schena (1999), supra).
Suggested substrates include silicon, silica, glass slides, glass chips, and silicon wafers. Alternatively, a procedure analogous to a dot or slot blot may also be used to arrange and link elements to the surface of a substrate using thermal, UV, chemical, or mechanical bonding procedures.
A typical array may be produced using available methods and machines well known to those of ordinary skill in the art and may contain any appropriate number of elements: (See, e.g., Schena, M. et al.
(1995) Science 270:467-470; Shalom D. et al. (1996) Genome Res. 6:639-645; Marshall, A. and J. Hodgson (1998) Nat. Biotechnol. 16:27-31.) Full length cDNAs, Expressed Sequence Tags (ESTs), or fragments or oligomers thereof may comprise the elements of the microarray. Fragments or oligomers suitable for hybridization can be selected using software well known in the art such as LASERGENE software (DNASTAR). The array elements are hybridized with polynucleotides in a biological sample. The polynucleotides in the biological sample are conjugated to a fluorescent label or other molecular tag for ease of detection.

After hybridization, nonhybridized nucleotides from the biological sample are removed, and a fluorescence scanner is used to detect hybridization at each array element.
Alternatively, laser desorbtion and mass spectrometry may be used for detection of hybridization.
The degree of complementarity and the relative abundance of each polynucleotide which hybridizes to an element on the microarray may be assessed. In one embodiment, microarray preparation and usage is described in detail below.
Tissue or Cell Sample Preparation Total RNA is isolated from tissue samples using the guanidinium thiocyanate method and poly(A)+ RNA is purified using the oligo-(dT) cellulose method. Each poly(A)+
RNA sample is reverse transcribed using MMLV reverse-transcriptase, 0.05 pg/~1 oligo-(dT) primer (2lmer), 1X first strand buffer, 0.03 units/~1 RNase inhibitor, 500 ~M dATP, 500 ~M dGTP, 500 ~M
dTTP, 40 ~M
dCTP, 40 ~M dCTP-Cy3 (BDS) or dCTP-Cy5 (Amersham Pharmacia Biotech). The reverse transcription reaction is performed in a 25 ml volume containing 200 ng poly(A)+ RNA with GEMBRIGHT kits (Incyte). Specific control poly(A)~ RNAs are synthesized by in vitro transcription from non-coding yeast genomic DNA. After incubation at 37° C for 2 hr, each reaction sample (one with Cy3 and another with Cy5 labeling) is treated with 2.5 ml of 0.5M sodium hydroxide and incubated for 20 minutes at 85° C to the stop the reaction and degrade the RNA. Samples are purified using two successive CHROMA SPIN 30 gel filtration spin columns (CLONTECH
Laboratories, Inc.
(CLONTECH), Palo Alto CA) and after combining, both reaction samples are ethanol precipitated using 1 ml of glycogen (1 mg/ml), 60 ml sodium acetate, and 300 ml of 100%
ethanol. The sample is then dried to completion using a SpeedVAC (Savant Instruments Inc., Holbrook NY) and resuspended in 14 ~l 5X SSC/0.2% SDS.
Microarray Preparation Sequences of the present invention are used to generate array elements. Each array element is amplified from bacterial cells containing vectors with cloned cDNA inserts.
PCR amplification uses primers complementary to the vector sequences flanking the cDNA insert. Array elements are amplified in thirty cycles of PCR from an initial quantity of 1-2 ng to a final quantity greater than 5 pg.
Amplified array elements are then purified using SEPHACRYL-400 (Amersham Pharmacia Biotech).
Purified array elements are immobilized on polymer-coated glass slides. Glass microscope slides (Corning) are cleaned by ultrasound in 0.1 % SDS and acetone, with extensive distilled water washes between and after treatments. Glass slides are etched in 4%
hydrofluoric acid (VWR
Scientific Products Corporation (VWR), West Chester PA), washed extensively in distilled water, and coated with 0.05% aminopropyl silane (Sigma) in 95% ethanol. Coated slides are cured in a 110°C

oven.
Array elements are applied to the coated glass substrate using a procedure described in US
Patent No. 5,807,522, incorporated herein by reference. 1 ~ 1 of the array element DNA, at an average concentration of 100 ngl~l, is loaded into the open capillary printing element by a high-speed robotic apparatus. The apparatus then deposits about 5 nl of array element sample per slide.
Microarrays are UV-crosslinked using a STRATALINKER UV-crosslinker (Stratagene).
Microarrays are washed at room temperature once in 0.2% SDS and three times in distilled water.
Non-specific binding sites are blocked by incubation of microarrays in 0.2%
casein in phosphate buffered saline (PBS) (Tropix, Inc., Bedford MA) for 30 minutes at 60°
C followed by washes in 0.2%
SDS and distilled water as before.
Hybridization Hybridization reactions contain 9 ~1 of sample mixture consisting of 0.2 ~g each of Cy3 and Cy5 labeled cDNA synthesis products in 5X SSC, 0.2%o SDS hybridization buffer.
The sample mixture is heated to 65° C for 5 minutes and is aliquoted onto the microarray surface and covered with an 1.8 cm2 coverslip. The arrays are transferred to a waterproof chamber having a cavity just slightly larger than a microscope slide. The chamber is kept at 100% humidity internally by the addition of 140 ~1 of 5X SSC in a corner of the chamber. The chamber containing the arrays is incubated for about 6.5 hours at 60° C. The arrays are washed for 10 min at 45° C in a first wash buffer (1X SSC, 0.1 SDS), three times for 10 minutes each at 45° C in a second wash buffer (O.1X SSC), and dried.
Detection Reporter-labeled hybridization complexes are detected with a microscope equipped with an Innova 70 mixed gas 10 W laser (Coherent, Inc., Santa Clara CA) capable of generating spectral lines at 488 nm for excitation of Cy3 and at 632 nm for excitation of CyS. The excitation laser light is focused on the array using a 20X microscope objective (Nikon, Inc., Melville NY). The slide containing the array is placed on a computer-controlled X-Y stage on the microscope and raster-scanned past the objective. The 1.8 cm x 1.8 cm array used in the present example is scanned with a resolution of 20 micrometers.
In two separate scans, a mixed gas multiline laser excites the two fluorophores sequentially.
Emitted light is split, based on wavelength, into two photomultiplier tube detectors (PMT 81477, Hamamatsu Photonics Systems, Bridgewater NJ) corresponding to the two fluorophores. Appropriate filters positioned between the array and the photomultiplier tubes are used to filter the signals. The emission maxima of the fluorophores used are 565 nm for Cy3 and 650 nm for CyS. Each array is typically scanned twice, one scan per fluorophore using the appropriate filters at the laser source, although the apparatus is capable of recording the spectra from both fluorophores simultaneously.
The sensitivity of the scans is typically calibrated using the signal intensity generated by a cDNA control species added to the sample mixture at a known concentration. A
specific location on the array contains a complementary DNA sequence, allowing the intensity of the signal at that location to be correlated with a weight ratio of hybridizing species of 1:100,000. When two samples from different sources (e.g., representing test and control cells), each labeled with a different fluorophore, are hybridized to a single array for the purpose of identifying genes that are differentially expressed, the calibration is done by labeling samples of the calibrating cDNA with the two fluorophores and adding identical amounts of each to the hybridization mixture.
The output of the photomultiplier tube is digitized using a 12-bit RTI-835H
analog-to-digital (A/D) conversion board (Analog Devices, Inc., Norwood MA) installed in an IBM-compatible PC
computer. The digitized data are displayed as an image where the signal intensity is mapped using a linear 20-color transformation to a pseudocolor scale ranging from blue (low signal) to red (high signal). The data is also analyzed quantitatively. Where two different fluorophores are excited and measured simultaneously, the data are first corrected for optical crosstalk (due to overlapping emission spectra) between the fluorophores using each fluorophore's emission spectrum.
A grid is superimposed over the fluorescence signal image such that the signal from each spot is centered in each element of the grid. The fluorescence signal within each element is then integrated to obtain a numerical value corresponding to the average intensity of the signal. The software used for signal analysis is the GEMTOOLS gene expression analysis program (Incyte).
XI. Complementary Polynucleotides Sequences complementary to the PKIN-encoding sequences, or any parts thereof, are used to detect, decrease, or inhibit expression of naturally occurring PKIN. Although use of oligonucleotides comprising from about 15 to 30 base pairs is described, essentially the same procedure is used with smaller or with larger sequence fragments. Appropriate oligonucleotides are designed using OLIGO
4.06 software (National Biosciences) and the coding sequence of PKIN. To inhibit transcription, a complementary oligonucleotide is designed from the most unique 5'sequence and used to prevent promoter binding to the coding sequence. To inhibit translation, a complementary oligonucleotide is designed to prevent ribosomal binding to the PKIN-encoding transcript.
XII. Expression of PKIN
Expression and purification of PI~IN is achieved using bacterial or virus-based expression systems. For expression of PI~IN in bacteria, cDNA is subcloned into an appropriate vector containing an antibiotic resistance gene and an inducible promoter that directs high levels of cDNA

transcription. Examples of such promoters include, but are not limited to, the trp-lac (tac) hybrid promoter and the TS or T7 bacteriophage promoter in conjunction with the lac operator regulatory element. Recombinant vectors are transformed into suitable bacterial hosts, e.g., BL21 (DE3).
Antibiotic resistant bacteria express PKIN upon induction with isopropyl beta-D-thiogalactopyranoside (IPTG). Expression of PKIN in eukaryotic cells is achieved by infecting insect or mammalian cell lines with recombinant Auto~raphica californica nuclear polyhedrosis virus (AcMNPV), commonly known as baculovirus. The nonessential polyhedrin gene of baculovirus is replaced with cDNA
encoding PKIN by either homologous recombination or bacterial-mediated transposition involving transfer plasmid intermediates. Viral infectivity is maintained and the strong polyhedrin promoter drives high levels of cDNA transcription. Recombinant baculovirus is used to infect Spodoptera fru~iperda (Sf9) insect cells in most cases, or human hepatocytes, in some cases. Infection of the latter requires additional genetic modifications to baculovirus. (See Engelhard, E.K. et al. (1994) Proc.
Natl. Acad. Sci. USA 91:3224-3227; Sandig, V. et al. (1996) Hum. Gene Ther.
7:1937-1945.) In most expression systems, PKIN is synthesized as a fusion protein with, e.g., glutathione S-transferase (GST) or a peptide epitope tag, such as FLAG or 6-His, permitting rapid, single-step, affinity-based purification of recombinant fusion protein. from crude cell lysates. GST, a 26-kilodalton enzyme from Schistosoma japonicum, enables the purification of fusion proteins on immobilized glutathione under conditions that maintain protein activity and antigenicity (Amersham Pharmacia Biotech). Following purification, the GST moiety can be proteolytically cleaved from PKIN at specifically engineered sites. FLAG, an 8-amino acid peptide, enables immunoaffinity purification using commercially available monoclonal and polyclonal anti-FLAG antibodies (Eastman Kodak). 6-His, a stretch of six consecutive histidine residues, enables purification on metal-chelate resins (QIAGEN). Methods for protein expression and purification are discussed in Ausubel (1995, supra, ch. 10 and 16). Purified PKIN obtained by these methods can be used directly in the assays shown in Examples XVI, XVII, XVIII, and XIX where applicable.
XIII. Functional Assays PKIN fiuiction is assessed by expressing the sequences encoding PKIN at physiologically elevated levels in mammalian cell culture systems. cDNA is subcloned into a mammalian expression vector containing a strong promoter that drives high levels of cDNA
expression. Vectors of choice include PCMV SPORT (Life Technologies) and PCR3.1 (Invitrogen, Carlsbad CA), both of which contain the cytomegalovirus promoter. 5-10 ,ug of recombinant vector are transiently transfected into a human cell line, for example, an endothelial or hematopoietic cell line, using either liposome formulations or electroporation. 1-2 ~g of an additional plasmid containing sequences encoding a marker protein are co-transfected. Expression of a marker protein provides a means to distinguish transfected cells from nontransfected cells and is a reliable predictor of cDNA expression from the recombinant vector. Marker proteins of choice include, e.g., Green Fluorescent Protein (GFP;
Clontech), CD64, or a CD64-GFP fusion protein. Flow cytometry (FCM), an automated, laser optics-based technique, is used to identify transfected cells expressing GFP or CD64-GFP and to evaluate the apoptotic state of the cells and other cellular properties. FCM detects and quantifies the uptake of fluorescent molecules that diagnose events preceding or coincident with cell death. These events include changes in nuclear DNA content as measured by staining of DNA with propidium iodide;
changes in cell size and granularity as measured by forward light scatter and 90 degree side light scatter; down-regulation of DNA synthesis as measured by decrease in bromodeoxyuridine uptake;
alterations in expression of cell surface and intracellular proteins as measured by reactivity with specific antibodies; and alterations in plasma membrane composition as measured by the binding of fluorescein-conjugated Annexin V protein to the cell surface. Methods in flow cytometry are discussed in Ormerod, M.G. (1994) Flow C, ometry, Oxford, New York NY.
The influence of PI~IN on gene expression can be assessed using highly purified populations of cells transfected with sequences encoding PKIN and either CD64 or CD64-GFP.
CD64 and CD64-GFP are expressed on the surface of transfected cells and bind to conserved regions of human immunoglobulin G (IgG). Transfected cells are e~ciently separated from nontransfected cells using magnetic beads coated with either human IgG or antibody against CD64 (DYNAL, Lake Success NY). mRNA can be purified from the cells using methods well known by those of skill in the art.
Expression of mRNA encoding PKIN and other genes of interest can be analyzed by northern analysis or microarray techniques.
XIV. Production of PKIN Specific Antibodies PKIN substantially purified using polyacrylamide gel electrophoresis (PAGE;
see, e.g., Harrington, M.G. (1990) Methods Enzymol. 182:488-495), or other purification techniques, is used to immunize rabbits and to produce antibodies using standard protocols.
Alternatively, the PKIN amino acid sequence is analyzed using LASERGENE
software (DNASTAR) to determine regions of high immunogenicity, and a corresponding oligopeptide is synthesized and used to raise antibodies by means known to those of skill in the art. Methods for selection of appropriate epitopes, such as those near the C-terminus or in hydrophilic regions are well described in the art. (See, e.g., Ausubel, 1995, supra, ch. 1 l.) Typically, oligopeptides of about 15 residues in length are synthesized using an ABI 431A
peptide synthesizer (Applied Biosystems) using FMOC chemistry and coupled to KI,H (Sigma-Aldrich, St. Louis MO) by reaction with N-maleimidobenzoyl-N-hydroxysuccinimide ester (MBS) to increase immunogenicity. (See, e.g., Ausubel, 1995, su ra.) Rabbits are immunized with the oligopeptide-KLH complex in complete Freund's adjuvant. Resulting antisera are tested for antipeptide and anti-PKIN activity by, for example, binding the peptide or PKIN to a substrate, blocking with 1 % BSA, reacting with rabbit antisera, washing, and reacting with radio-iodinated goat anti-rabbit IgG.
XV. Purification of Naturally Occurring PKIN Using Specific Antibodies Naturally occurring or recombinant PKIN is substantially purified by immunoaffinity chromatography using antibodies specific for PKIN. An immunoaffinity column is constructed by covalently coupling anti-PKIN antibody to an activated chromatographic resin, such as CNBr-activated SEPHAROSE (Amersham Pharmacia Biotech). After the coupling, the resin is blocked and washed according to the manufacturer's instructions.
Media containing PKIN are passed over the immunoaffinity column, and the column is washed under conditions that allow the preferential absorbance of PK1N (e.g., high ionic strength buffers in the presence of detergent). The column is eluted under conditions that disrupt antibody/PKIN binding (e.g., a buffer of pH 2 to pH 3, or a high concentration of a chaotrope, such. as urea or thiocyanate ion), and PK1N is collected.
XVI. Identification of Molecules Which Interact with PKIN
PKIN, or biologically active fragments thereof, are labeled with'ZSI Bolton-Hunter reagent.
(See, e.g., Bolton A.E. and W.M. Hunter (1973) Biochem. J. 133:529-539.) Candidate molecules previously arrayed in the wells of a multi-well plate are incubated with the labeled PK1N, washed, and any wells with labeled PKIN complex are assayed. Data obtained using different concentrations of PKIN are used to calculate values for the number, affinity, and association of PKIN with the candidate molecules.
Alternatively, molecules interacting with PK1N are analyzed using the yeast two-hybrid system as described in Fields, S. and O. Song (1989) Nature 340:245-246, or using commercially available kits based on the two-hybrid system, such as the MATCHMAKER system (Clontech).
PK1N may also be used in the PATHCALLING process (CuraGen Corp., New Haven CT) which employs the yeast two-hybrid system in a high-throughput manner to determine all interactions between the proteins encoded by two large libraries of genes (Nandabalan, K.
et al. (2000) U.S.
Patent No. 6,057,101).
XVII. Demonstration of PKIN Activity Generally, protein kinase activity is measured by quantifying the phosphorylation of a protein substrate by PKIN in the presence of [y 32P]ATP. PKIN is incubated with the protein substrate, sap-ATP, and an appropriate kinase buffer. The 32P incorporated into the substrate is separated from free 32P-ATP by electrophoresis and the incorporated 32P is counted using a radioisotope counter.
The amount of incorporated 32P is proportional to the activity of PKIN. A
determination of the specific amino acid residue phosphorylated is made by phosphoamino acid analysis of the hydrolyzed protein.
In one alternative, protein kinase activity is measured by quantifying the transfer of gamma phosphate from adenosine triphosphate (ATP) to a serine, threonine or tyrosine residue in a protein substrate. The reaction occurs between a protein kinase sample with a biotinylated peptide substrate and gamma 32P-ATP. Following the reaction, free avidin in solution is added for binding to the biotinylated 32P-peptide product. The binding sample then undergoes a centrifugal ultrafiltration process with a membrane which will retain the product-avidin complex and allow passage of free gamma 32P-ATP. The reservoir of the centrifuged unit containing the 32P-peptide product as retentate is then counted in a scintillation counter. This procedure allows assay of any type of protein kinase sample, depending on the peptide substrate and kinase reaction buffer selected. This assay is provided in kit form (ASUA, Affinity Ultrafiltration Separation Assay, Transbio Corporation, Baltimore MD, U.S. Patent No. 5,869,275). Suggested substrates and their respective enzymes include but are not limited to: Histone H1 (Sigma) and p34~a°2kinase, Annexin I, Angiotensin (Sigma) and EGF receptor kinase, Annexin II and src kinase, ERKl & ERK2 substrates and MEK, and myelin basic protein and ERK (Pearson, J.D. et al. (1991) Methods Enzymol. 200:62-81).
In another alternative, protein kinase activity of PKIN is demonstrated in an assay containing PKIN, 50.1 of kinase buffer, 1 ~.g substrate, such as myelin basic protein (MBP) or synthetic peptide substrates, 1 mM DTT, 10 ~,g ATP, and 0.5 ~.Ci [y 3'P]ATP. The reaction is incubated at 30°C for minutes and stopped by pipetting onto P81 paper. The unincorporated [y 32P]ATP
is removed by 25 washing and the incorporated radioactivity is measured using a scintillation counter. Alternatively, the reaction is stopped by heating to 100°C in the presence of SDS loading buffer and resolved on a 12%
SDS polyacrylamide gel followed by autoradiography. The amount of incorporated 32P is proportional to the activity of PKIN.
In yet another alternative, adenylate kinase or guanylate kinase activity may be measured by 30 the incorporation of 32P from [y-32P]ATP into ADP or GDP using a gamma radioisotope counter.
The enzyme, in a kinase buffer, is incubated together with the appropriate nucleotide mono-phosphate substrate (AMP or GMP) and 32P-labeled ATP as the phosphate donor. The reaction is incubated at 37°C and terminated by addition of trichloroacetic acid. The acid extract is neutralized and subjected to gel electrophoresis to separate the mono-, di-, and triphosphonucleotide fractions. The diphosphonucleotide fraction is excised and counted. The radioactivity recovered is proportional to the enzyme activity.
In yet another alternative, other assays for PKIN include scintillation proximity assays (SPA), scintillation plate technology and filter binding assays. Useful substrates include recombinant proteins tagged with glutathione transferase, or synthetic peptide substrates tagged with biotin. Inhibitors of PKIN activity, such as small organic molecules, proteins or peptides, may be identified by such assays.
XVIII. Enhancement/Inhibition of Protein Kinase Activity Agonists or antagonists of PKIN activation or inhibition may be tested using assays described in section XVII. Agonists cause an increase in PKIN activity and antagonists cause a decrease in PKIN activity.
XIX. Kinase Binding Assay Binding of PKIN to a FLAG-CD44 cyt fusion protein can be determined by incubating PKIN
to anti-PKIN-conjugated immunoaffinity beads followed by incubating portions of the beads (having 10-20 ng of protein) with 0.5 ml of a binding buffer (20 mM Tris-HCL (pH 7.4), 150 mM NaCl, 0.1 %
bovine serum albumin, and 0.05% Triton X-100) in the presence of lzsl-labeled FLAG-CD44cyt fusion protein (5,000 cpm/ng protein ) at 4 °C for 5 hours. Following binding, beads were washed thoroughly in the binding buffer and the bead-bound radioactivity measured in a scintillation counter (Bourguignon, L.Y.W. et al. (2001) J. Biol. Chem. 276:7327-7336). The amount of incorporated 32P is proportional to the amount of bound PKIN.
Various modifications and variations of the described methods and systems of the invention will be apparent to those skilled in the art without departing from the scope and spirit of the invention.
Although the invention has been described in connection with certain embodiments, it should be understood that the invention as claimed should not be unduly limited to such specific embodiments.
Indeed, various modifications of the described modes for carrying out the invention which are obvious to those skilled in molecular biology or related fields are intended to be within the scope of the following claims.

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<110> INCYTE GENOMICS, INC.
YUE, Henry KHAN, Farrah A.
GURURAJAN, Rajagopal HAFALIA, April J. A.
WALIA, Narinder K.
PATTERSON, Chandra RAMKUMAR, Jayalaxini GANDHI, Ameena R.
POLICKY, Jennifer L.
BAUGHN, Mariah R.
TRIBOULEY, Catherine M.
THORNTON, Michael BANDMAN, Olga NGUYEN, Danniel B.
LU, Yan BURFORD, Neil LAL, Preeti DING, Li YAO, Monique G.
ELLIOTT, Vicki S.
RECIPON, Shirley A.
KEARNEY, Llam LU, Dyung Aina M.
GREENWALD, Sara R.
TANG, Y. Tom XU, Yuming WALSH, Roderick T.
GIETZEN, Kimberly J.
YANG, Junming HILLMAN, Jennifer L.
<120> HUMAN KINASES
<130> PI-0162 PCT
<140> To Be Assigned <141> Herewith <150> 60/220,038; 60/222,112; 60/222,831; 60/224,729 <151> 2000-07-21; 2000-07-28; 2000-08-04; 2000-08-11 <160> 40 <170> PERL Program <210> 1 <211> 1297 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 2564295CD1 <400> 1 Met Ala Val Pro Ser Leu Trp Pro Trp Gly Ala Cys Leu Pro Val Pro Ser Leu Asp Ile Arg Ser Glu Val Ala Glu Leu Arg G1n Leu Glu Asn Cys Ser Val Val Glu Gly His Leu Gln Ile Leu Leu Met Phe Thr Ala Thr Gly Glu Asp Phe Arg Gly Leu Ser Phe Pro Arg Leu Thr Gln Val Thr Asp Tyr Leu Leu Leu Phe Arg Val Tyr Gly Leu Glu Ser Leu Arg Asp Leu Phe Pro Asn Leu Ala Val Ile Arg Gly Thr Arg Leu Phe Leu Gly Tyr Ala Leu Val Ile Phe Glu Met Pro His Leu Arg Asp Va1 Ala Leu Pro Ala Leu Gly Ala Val Leu Arg Gly Ala Val Arg Val Glu Lys Asn Gln Glu Leu Cys His Leu Ser Thr Ile Asp Trp Gly Leu Leu Gln Pro Ala Pro Gly Ala Asn His Ile Val Gly Asn Lys Leu Gly Glu Glu Cys Ala Asp Val Cys Pro G1y Val Leu Gly Ala Ala Gly G1u Pro Cys Ala Lys Thr Thr Phe Ser Gly His Thr Asp Tyr Arg Cys Trp Thr Ser Ser His Cys Gln Arg Val Cys Pro Cys Pro His Gly Met Ala Cys Thr Ala Arg Gly Glu Cys Cys His Thr Glu Cys Leu Gly Gly Cys Ser Gln Pro Glu Asp Pro Arg Ala Cys Val Ala Cys Arg His Leu Tyr Phe Gln Gly Ala Cys Leu Trp Ala Cys Pro Pro Gly Thr Tyr Gln Tyr Glu Ser Trp Arg Cys Val Thr Ala Glu Arg Cys Ala Ser Leu His Ser Val Pro Gly Arg Ala Ser Thr Phe Gly Ile His Gln G1y Ser Cys 290 , 295 300 Leu Ala Gln Cys Pro Ser Gly Phe Thr Arg Asn Ser Ser Ser Ile Phe Cys His Lys Cys Glu Gly Leu Cys Pro Lys Glu Cys Lys Val Gly Thr Lys Thr Ile Asp Ser Ile Gln Ala Ala Gln Asp Leu Val Gly Cys Thr His Val Glu Gly Ser Leu Ile Leu Asn Leu Arg Gln Gly Tyr Asn Leu Glu Pro Gln Leu Gln His Ser Leu Gly Leu Val Glu Thr Ile Thr Gly Phe Leu Lys Ile Lys His Ser Phe Ala Leu Val Ser Leu Gly Phe Phe Lys Asn Leu Lys Leu Ile Arg Gly Asp Ala Met Val Asp Gly Asn Tyr Thr Leu Tyr Val Leu Asp Asn Gln Asn Leu Gln Gln Leu G1y Ser Trp Val Ala Ala Gly Leu Thr Ile Pro Val Gly Lys Ile Tyr Phe Ala Phe Asn Pro Arg Leu Cys Leu Glu His Ile Tyr Arg Leu Glu Glu Val Thr Gly Thr Arg Gly Arg G1n Asn Lys Ala Glu Ile Asn Pro Arg Thr Asn Gly Asp Arg Ala Ala Cys Gln Thr Arg Thr Leu Arg Phe Val Ser Asn Val Thr Glu 485 . 490 495 Ala Asp Arg Ile Leu Leu Arg Trp Glu Arg Tyr Glu Pro Leu Glu Ala Arg Asp Leu Leu Ser Phe Ile Val Tyr Tyr Lys Glu Ser Pro Phe Gln Asn Ala Thr Glu His Val Gly Pro Asp Ala Cys Gly Thr Gln Ser Trp Asn Leu Leu Asp Val Glu Leu Pro Leu Ser Arg Thr Gln Glu Pro Gly Val Thr Leu Ala Ser Leu Lys Pro Trp Thr Gln Tyr Ala Val Phe Val Arg Ala Ile Thr Leu Thr Thr Glu Glu Asp Ser Pro His Gln Gly Ala Gln Ser Pro Ile Val Tyr Leu Arg Thr Leu Pro Ala Ala Pro Thr Va1 Pro Gln Asp Val Ile Ser Thr Ser Asn Ser Ser Ser His Leu Leu Val Arg Trp Lys Pro Pro Thr Gln Arg Asn Gly Asn Leu Thr Tyr Tyr Leu Va1 Leu Trp Gln Arg Leu Ala Glu Asp Gly Asp Leu Tyr Leu Asn Asp Tyr Cys His Arg Gly Leu Arg Leu Pro Thr Ser Asn Asn Asp Pro Arg Phe Asp Gly Glu Asp Gly Asp Pro Glu Ala Glu Met Glu Ser Asp Cys Cys Pro Cys Gln His Pro Pro Pro Gly Gln Val Leu Pro Pro Leu Glu Ala Gln Glu Ala Ser Phe Gln Lys Lys Phe Glu Asn Phe Leu His Asn Ala Ile Thr Ile Pro Ile Ser Pro Trp Lys Val Thr Ser Ile Asn Lys Ser Pro Gln Arg Asp Ser Gly Arg His Arg Arg Ala Ala Gly Pro Leu Arg Leu Gly Gly Asn Ser Ser Asp Phe Glu Ile Gln Glu Asp Lys Val Pro Arg Glu Arg Ala Val Leu Ser Gly Leu Arg His Phe Thr Glu Tyr Arg I1e Asp Ile His Ala Cys Asn His Ala Ala His Thr Val Gly Cys Ser Ala Ala Thr Phe Val Phe Ala Arg Thr Met Pro His Arg Glu Ala Asp Gly Ile Pro Gly Lys Val Ala Trp Glu Ala Ser Ser Lys Asn Ser Val Leu Leu Arg Trp Leu Glu Pro Pro Asp Pro Asn Gly Leu Ile Leu Lys Tyr Glu Ile Lys Tyr Arg Arg Leu Gly Glu Glu Ala Thr Val Leu Cys Val Ser Arg Leu Arg Tyr Ala Lys Phe Gly Gly Val His Leu Ala Leu Leu Pro Pro Gly Asn Tyr Ser A1a Arg Val Arg Ala Thr Ser Leu Ala Gly Asn Gly Ser Trp Thr Asp Ser Val Ala Phe Tyr Ile Leu Gly Pro Glu Glu Glu Asp Ala Gly Gly Leu His Val Leu Leu Thr Ala Thr Pro Val Gly Leu Thr Leu Leu Ile Val Leu Ala Ala Leu Gly Phe Phe Tyr Gly Lys Lys Arg Asn Arg Thr Leu Tyr Ala Ser Val Asn Pro Glu Tyr Phe Ser Ala Ser Asp Met Tyr Val Pro Asp Glu Trp Glu Val Pro Arg Glu Gln Ile Ser Ile Ile Arg Glu Leu Gly Gln Gly Ser Phe Gly Met Val Tyr Glu Gly Leu Ala Arg Gly Leu Glu Ala Gly Glu Glu Ser Thr Pro Val Ala Leu Lys Thr Val Asn Glu Leu Ala Ser Pro Arg Glu Cys Ile Glu Phe Leu Lys Glu Ala Ser Val Met Lys Ala Phe Lys Cys His His Val Val Arg Leu Leu Gly Val Val Ser G1n Gly Gln Pro Thr Leu Val Ile Met Glu Leu Met Thr Arg Gly Asp Leu Lys Ser His Leu Arg Ser Leu Arg Pro Glu Ala Glu Asn Asn Pro Gly Leu Pro Gln Pro Ala Leu Gly Glu Met Ile Gln Met Ala Gly Glu I1e Ala Asp G1y Met Ala Tyr Leu Ala Ala Asn Lys Phe Val His Arg Asp Leu Ala Ala Arg Asn Cys Met Val Ser Gln Asp Phe Thr Val Lys Ile Gly Asp Phe Gly Met Thr Arg Asp Val Tyr Glu Thr Asp Tyr Tyr Arg Lys Gly Gly Lys Gly Leu Leu Pro Val Arg Trp Met Ala Pro Glu Ser Leu Lys Asp Gly Ile Phe Thr Thr His Ser Asp Val Trp Ser Phe Gly Val Val Leu Trp Glu Ile Val Thr Leu Ala Glu Gln Pro Tyr Gln Gly Leu Ser Asn Glu Gln ' 1190 1195 1200 Val Leu Lys Phe Val Met Asp Gly Gly Val Leu Glu Glu Leu Glu Gly Cys Pro Leu Gln Leu Gln Glu Leu Met Ser Arg Cys Trp Gln ' 1220 1225 1230 Pro Asn Pro Arg Leu Arg Pro Ser Phe Thr His Ile Leu Asp Ser Ile Gln Glu Glu Leu Arg Pro Ser Phe Arg Leu Leu Ser Phe Tyr Tyr Ser Pro G1u Cys Arg Gly Ala Arg Gly Ser Leu Pro Thr Thr Asp Ala Glu Pro Asp Ser Ser Pro Thr Pro Arg Asp Cys Ser Pro Gln Asn Gly Gly Pro Gly His <210> 2 <211> 718 <212> PRT
<213> Homo Sapiens <220>
<221> misc_teature <223> Incyte ID No: 2837050CD1 <400> 2 Met Met Glu Glu Leu His Ser Leu Asp Pro Arg Arg G1n Glu Leu Leu Glu Ala Arg Phe Thr Arg Val Gly Val Ser Lys Gly Pro Leu Asn Ser Glu Ser Ser Asn Gln Ser Leu Cys Ser Val Gly Ser Leu Ser Asp Lys Glu Val Glu Thr Pro Glu Lys Lys Gln Asn Asp Gln Arg Asn Arg Lys Arg Lys Ala Glu Pro Tyr Glu Thr Ser Gln Gly Lys Gly Thr Pro Arg Gly His Lys Ile Ser Asp Tyr Phe Glu Arg Arg Val G1u Gln Pro Leu Tyr Gly Leu Asp G1y Ser Ala Ala Lys Glu Ala Thr Glu Glu Gln Ser Ala Leu Pro Thr Leu Met Ser Val Met Leu Ala Lys Pro Arg Leu Asp Thr Glu His Val A1a G1n Arg Gly Ala Gly Leu Cys Phe Thr Phe Val Ser Ala Gln Gln Asn Ser Pro Ser Ser Thr Gly Ser Gly Asn Thr Glu His Ser Cys Ser Ser Gln Lys Gln Ile Ser Ile Gln His Arg Gln Thr Gln Ser As_p Leu Thr Ile Glu Lys Ile Ser Ala Leu Glu Asn Ser Lys Asn Ser Asp Leu Glu Lys Lys Glu Gly Arg Ile Asp Asp Leu Leu Arg Ala Asn Cys Asp Leu Arg Arg Gln Ile Asp Glu Gln Gln Lys Met Leu Glu Lys Tyr Lys Glu Arg Leu Asn Arg Cys Val Thr Met Ser Lys Lys Leu Leu Ile Glu Lys Ser Lys Gln Glu Lys Met Ala Cys Arg Asp Lys Ser Met Gln Asp Arg Leu Arg Leu Gly His Phe Thr Thr Val Arg His Gly Ala Ser Phe Thr Glu Gln Trp Thr Asp Gly Tyr Ala Phe Gln Asn Leu Ile Lys Gln Gln Glu Arg Ile Asn Ser Gln Arg Glu Glu Ile Glu Arg Gln Arg Lys Met Leu Ala Lys Arg Lys Pro Pro Ala Met Gly Gln Ala Pro Pro Ala Thr Asn Glu Gln Lys Gln Arg Lys Ser Lys Thr Asn Gly Ala Glu Asn Glu Thr Leu Thr Leu Ala Glu Tyr His Glu Gln Glu Glu Ile Phe Lys Leu Arg Leu Gly 350 , 355 360 His Leu Lys Lys Glu Glu Ala Glu Ile Gln Ala Glu Leu Glu Arg Leu Glu Arg Va1 Arg Asn Leu His Ile Arg Glu Leu Lys Arg Ile His Asn Glu Asp Asn Ser Gln Phe Lys Asp His Pro Thr Leu Asn Asp Arg Tyr Leu Leu Leu His Leu Leu Gly Arg Gly Gly Phe Ser Glu Val Tyr Lys Ala Phe Asp Leu Thr Glu G1n Arg Tyr Val Ala Val Lys Ile His Gln Leu Asn Lys Asn Trp Arg Asp Glu Lys Lys Glu Asn Tyr His Lys His Ala Cys Arg Glu Tyr Arg Ile His Lys Glu Leu Asp His Pro Arg Ile Val Lys Leu Tyr Asp Tyr Phe Ser Leu Asp Thr Asp Ser Phe Cys Thr Val Leu Glu Tyr Cys Glu Gly Asn Asp Leu Asp Phe Tyr Leu Lys Gln His Lys Leu Met Ser Glu Lys Glu Ala Trp Ser I1e Ile Met Gln I1e Val Asn A1a Leu Lys Tyr Leu Asn Glu Ile Lys Pro Pro Ile Ile His Tyr Asp Leu Lys Pro Gly Asn Ile Leu Leu Val Asn Gly Thr Val Cys Gly Glu Arg Lys Ile Thr Asp Phe Gly Leu Ser Lys Ile Met Asp Asp Asp Ser Tyr Asn Ser Va1 Gly Gly Met Glu Leu Thr Ser Gln Gly Ala G1y Thr Tyr Trp Tyr Leu Pro Pro Glu Cys Phe Val Val Glu Lys Glu Pro Pro Lys Ile Ser Asn Lys Val Asp Val Trp Ser Val Gly Val 605 6l0 615 Ile Phe Tyr Gln Cys Leu Ser Gly Gly Lys Pro Phe Gly His Asn Gln Ser Gln Gln Asp Ile Leu G1n Glu Asn Thr Ile Leu Lys Ala Ala Glu Val Gln Phe Pro Pro Lys Pro Val Val Thr Pro Glu Ala Lys Ala Phe Ile Arg Arg Cys Leu Ala Tyr Arg Lys Glu Asp Cys Ile Asp Ala Gln Gln Leu Ala Cys Asp Pro Tyr Leu Leu Pro His Ile Arg Lys Ser Val Ser Thr Ser Ser Pro Ala Gly Ala Ala Ile Ala Ser Thr Ser Gly Ala Ser Asn Asn Ser Ser Ser Asn <210> 3 <211> 497 <212> PRT
<213> Homo Sapiens <220>
<221> mist feature <223> Incyte ID No: 7474590CD1 <400> 3 Met Tyr Ser Asp Ser G1u Asp Glu Ser Ser Glu Leu Ser Thr Val Leu Ser Met Phe Glu Glu Lys Glu Phe Thr Arg Gln Tyr Thr Val Leu Lys Thr Leu Ser Gln His Gly Thr Thr Glu Val Arg Leu Cys Ser His His Leu Thr Gly Val Thr Val Ala Val Lys Ala Leu Lys Tyr Gln Arg Trp Trp Glu Pro Lys Val Ser Glu Val Glu Ile Met Lys Met Leu Ser His Pro Asn I1e Val Ser Leu Leu Gln Val Tle Glu Thr Glu Gln Asn Ile Tyr Leu Ile Met Glu Val Ala Gln Gly Thr Gln Leu His Asn Arg Val Gln Glu Ala Arg Cys Leu Lys Glu Asp Glu Ala Arg Ser Ile Phe Val Gln Leu Leu Ser Ala I1e Gly Tyr Cys His Gly Glu Gly Val Val His Arg Asp Leu Lys Pro Asp Asn Val Tle Val Asp Glu His Gly Asn Val Lys Ile Val Asp Phe Gly Leu Gly Ala Arg Phe Met Pro Gly G1n Lys Leu Glu Arg Leu Cys Gly Ala Phe Gln Phe Ile Pro Pro Glu Ile Phe Leu Gly Leu Pro Tyr Asp Gly Pro Lys Val Asp Ile Trp Ala Leu Gly Val Leu Leu Tyr Tyr Met Val Thr Gly Ile Phe Pro Phe Val Gly Ser Thr Leu Ser Glu Ile Ser Lys Glu Val Leu Gln Gly Arg Tyr Glu Ile Pro Tyr Asn Leu Ser Lys Asp Leu Arg Ser Met Ile Gly Leu Leu Leu Ala Thr Asn Ala Arg Gln Arg Pro Thr Ala Gln Asp Leu Leu Ser His Pro Trp Leu Gln Glu Gly Glu Lys Thr Ile Thr Phe His Ser Asn Gly Asp Thr Ser Phe Pro Asp Pro Asp Ile Met Ala Ala Met Lys Asn TIe Gly Phe His Val Gln Asp Ile Arg GIu Ser Leu Lys His Arg Lys Phe Asp Glu Thr Met Ala Thr Tyr Asn Leu Leu Arg Ala Glu Ala Cys Gln Asp Asp Gly Asn Tyr Val Gln Thr Lys Leu Met Asn Pro Gly Met Pro Pro Phe Pro Ser Val Thr Asp Ser Gly Ala Phe Ser Leu Pro Pro Arg Arg Arg Ala Ser Glu Pro Ser Phe Lys Val Leu Val Ser Ser Thr Glu Glu His Gln Leu Arg Gln Thr Gly Gly Thr Asn Ala Pro Phe Pro Pro Lys Lys Thr Pro Thr Met Gly Arg Ser Gln Lys Gln Lys Arg Ala Met Thr Ala Pro Cys Ile Cys Leu Leu Arg Asn Thr Tyr Ile Asp Thr Glu Asp Ser Ser Phe Cys Thr Ser Ser Gln Ala Glu Lys Thr Ser Ser Asp Pro Glu Lys Ser Glu Thr Ser Thr Ser Cys Pro Leu Thr Pro Arg Gly Trp Arg Lys Trp Lys Lys Arg Ile Val Ala Cys Ile Gln Thr Leu Cys Cys Cys Thr Leu Pro Gln Lys Lys Cys Pro Arg Ser Val His Pro Gln Lys <210> 4 <211> 74l <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 7474594CD1 <400> 4 Met Ser Gly Leu Va1 Leu Met Leu Ala Ala Arg Cys Ile Val Gly Ser Ser Pro Leu Cys Arg Cys Arg Arg Arg Arg Pro Arg Arg Ile Gly Ala Gly Pro Gly Arg Asp Asp Pro Gly Arg Lys Ala Ala Ala Ala Gly Gly Ser Gly Ser Pro Asn Ala Ala Leu Ser Arg Pro Arg Pro Ala Pro Ala Pro Gly Asp Ala Pro Pro Arg Ala Ala A1a Ser Ala Ala Ala Ala Ala Ala Ala Ala Ala Gly Thr Glu Gln Va1 Asp Gly Pro Leu Arg Ala Gly Pro Ala Asp Thr Pro Pro Ser Gly Trp Arg Met Gln Cys Leu Ala Ala Ala Leu Lys Asp Glu Thr Asn Met Ser Gly Gly Gly G1u Gln Ala Asp Ile Leu Pro Ala Asn Tyr Val Val Lys Asp Arg Trp Lys Val Leu Lys Lys Ile G1y Gly Gly Gly Phe Gly Glu Ile Tyr Glu Ala Met Asp Leu Leu Thr Arg Glu Asn Val Ala Leu Lys Val Glu Ser Ala Gln Gln Pro Lys Gln Val Leu Lys Met Glu Val Ala Va1 Leu Lys Lys Leu Gln Gly Lys Asp His Val Cys Arg Phe Ile Gly Cys Gly Arg Asn Glu Lys Phe Asn Tyr Val Val Met G1n Leu Gln Gly Arg Asn Leu A1a Asp Leu Arg Arg Ser Gln Pro Arg G1y Thr Phe Thr Leu Ser Thr Thr Leu Arg Leu Gly Lys Gln Ile Leu Glu Ser Ile Glu Ala Ile His Ser Val Gly Phe Leu His Arg Asp Ile Lys Pro Ser Asn Phe Ala Met Gly Arg Leu Pro Ser Thr Tyr Arg Lys Cys Tyr Met Leu Asp Phe Gly Leu Ala Arg Gln Tyr Thr Asn Thr Thr Gly Asp Val Arg Pro Pro Arg Asn Val Ala Gly Phe Arg Gly Thr Va1 Arg Tyr Ala Ser Val Asn Ala His Lys Asn Arg Glu Met Gly Arg His Asp Asp Leu Trp Ser Leu Phe Tyr Met Leu Val Glu Phe Ala Val Gly Gln Leu Pro Trp Arg Lys I1e Lys Asp Lys Glu Gln Val Gly Met Ile Lys Glu Lys Tyr Glu His Arg Met Leu Leu Lys His Met Pro Ser Glu Phe His Leu Phe Leu Asp His Ile Ala Ser Leu Asp Tyr Phe Thr Lys Pro Asp Tyr Gln Leu Ile Met Ser Val Phe Glu Asn Ser Met Lys Glu Arg Gly Ile Ala Glu Asn Glu Ala Phe Asp Trp Glu Lys Ala Gly Thr Asp Ala Leu Leu Ser Thr Ser Thr Ser Thr Pro Pro G1n Gln Asn Thr Arg Gln Thr Ala Ala Met Phe Gly Val Va1 Asn Val Thr Pro Val Pro Gly Asp Leu Leu Arg Glu Asn Thr Glu Asp Val Leu Gln Gly Glu His Leu Ser Asp Gln Glu Asn Ala Pro Pro Ile Leu Pro Gly Arg Pro Ser Glu Gly Leu G1y Pro Ser Pro His Leu.Val Pro His Pro Gly Gly Pro Glu Ala G1u Val Trp Glu Glu Thr Asp Va1 Asn Arg Asn Lys Leu Arg Ile Asn Ile Gly Lys Val Thr Ala Ala Arg Ala Lys Gly Val Gly G1y Leu Phe Ser His Pro Arg Phe Pro Ala Leu Cys Pro Cys Pro Val Pro Pro Lys His Pro Val Pro Gly His Leu Pro Ala Cys Pro Ala Ser Val Ser Arg Ser Leu Pro Ala Leu Ala Ser Leu Cys Leu Pro Ser Ser Ser Ser Ser Val Ser Phe Thr Leu Arg Arg Pro Ser Ala His Ser Arg Leu Ile Ser Pro Ser Ser Trp His Ser Pro Leu Leu Gln Ser Pro Cys Val Glu Glu 605 ~ 610 615 Glu Gln Ser Arg Gly Met Gly Val Pro Ser Ser Pro Val Arg Ala Pro Pro Asp Ser Pro Thr Thr Pro Val Arg Ser Leu Arg Tyr Arg Arg Val Asn Ser Pro Glu Ser Glu Arg Leu Ser Thr Ala Asp G1y Arg Val Glu Leu Pro Glu Arg Arg Trp Val Trp Gly Gln Gly His Gly Trp Gly Pro Arg Pro Ser Pro Pro Ser Arg Gly Trp Ser Gly Gly Lys Val Arg Cys Val Ala Glu Val Gly Arg Pro Trp G1u Val Leu Arg Gly Leu Tyr Leu Gly Leu Gly Ser Asp Ser Val Gly Ala Arg Asp Arg Ala Trp G1u Asn Gl.n Trp Gly Ile Gln Arg Gly Pro Gly Ser Cys Gln Glu Thr <210> 5 <211> 645 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 7477585CD1 <400> 5 Met Leu Lys Phe Gln Glu Ala Ala Lys Cys Val Ser G1y Ser Thr A1a Ile Ser Thr Tyr Pro Lys Thr Leu Ile Ala Arg Arg Tyr Val Leu Gln Gln Lys Leu Gly Ser Gly Ser Phe Gly Thr Val Tyr Leu Val Ser Asp Lys Lys Ala Lys Arg Gly Glu Glu Leu Lys Val Leu 50 55 , 60 Lys Glu Ile Ser Val Gly Glu Leu Asn Pro Asn Glu Thr Val Gln Ala Asn Leu Glu Ala Gln Leu Leu Ser Lys Leu Asp His Pro Ala Ile Val Lys Phe His Ala Ser Phe Val Glu Gln Asp Asn Phe Cys Ile Ile Thr Glu Tyr Cys Glu Gly Arg Asp Leu Asp Asp Lys I1e Gln Glu Tyr Lys Gln Ala Gly Lys Ile Phe Pro Glu Asn Gln Ile Ile Glu Trp Phe Ile Gln Leu Leu Leu Gly Val Asp Tyr Met His Glu Arg Arg Ile Leu His Arg Asp Leu Lys Ser Lys Asn Val Phe Leu Lys Asn Asn Leu Leu Lys Ile Gly Asp Phe Gly Val Ser Arg Leu Leu Met Gly Ser Cys Asp Leu A1a Thr Thr Leu Thr Gly Thr Pro His Tyr Met Ser Pro G1u Ala Leu Lys His Gln Gly Tyr Asp Thr Lys Ser Asp Ile Trp Ser Leu Ala Cys Ile Leu Tyr Glu Met Cys Cys Met Asn His Ala Phe Ala Gly Ser Asn Phe Leu Ser Ile Val Leu Lys Ile Va1 Glu Gly Asp Thr Pro Ser Leu Pro Glu Arg Tyr Pro Lys Glu Leu Asn Ala Ile Met Glu Ser Met Leu Asn Lys Asn Pro Ser Leu Arg Pro Ser Ala Ile Glu Ile Leu Lys Ile Pro Tyr Leu Asp Glu Gln Leu Gln Asn Leu Met Cys Arg Tyr Ser G1u Met Thr Leu Glu Asp Lys Asn Leu Asp Cys Gln Lys Glu Ala Ala His Ile Ile Asn A1a Met Gln Lys Arg Ile His Leu Gln Thr Leu Arg Ala Leu Ser Glu Val Gln Lys Met Thr Pro Arg Glu Arg Met Arg Leu Arg Lys Leu Gln Ala Ala Asp Glu Lys Ala Arg Lys Leu Lys Lys Ile Va1 Glu Glu Lys Tyr Glu Glu Asn Ser Lys Arg Met Gln Glu Leu Arg Ser Arg Asn Phe Gln Gln Leu Ser Val Asp Val Leu His Glu Lys Thr His Leu Lys Gly Met Glu G1u Lys Glu Glu Gln Pro Glu Gly Arg Leu Ser Cys Ser Pro Gln Asp Glu Asp Glu Glu Arg Trp Gln Gly Arg Glu Glu Glu Ser Asp Glu Pro Thr Leu Glu Asn Leu Pro Glu Ser Gln Pro Ile Pro Ser Met Asp Leu His Glu Leu Glu Ser Ile Va1 Glu Asp Ala Thr Ser Asp Leu Gly Tyr His Glu I1e Pro Glu Asp Pro Leu Val Ala Glu Glu Tyr Tyr Ala Asp Ala Phe Asp Ser Tyr Cys Val Glu Ser Asp Glu Glu Glu Glu Glu Ile Ala Leu Glu Arg Pro Glu Lys Glu Ile Arg Asn Glu Gly Ser Gln Pro Ala Tyr Arg Thr Asn Gln Gln Asp Ser Asp Ile Glu Ala Leu Ala Arg Cys Leu Glu Asn Val Leu Gly Cys Thr Ser Leu Asp Thr Lys Thr Ile Thr Thr Met Ala Glu Asp Met Ser Pro Gly Pro Pro Ile Phe Asn Ser Val Met Ala Arg Thr Lys Met Lys Arg Met Arg Glu Ser Ala Met Gln Lys Leu Gly Thr Glu Val Phe Glu Glu Val Tyr Asn Tyr Leu Lys Arg Ala Arg His Gln Asn Ala Ser Glu Ala Glu Ile Arg Glu Cys Leu Glu Lys Va1 Val Pro Gln Ala Ser Asp Cys Phe Glu Val Asp Gln Leu Leu Tyr Phe Glu Glu Gln Leu Leu Ile Thr Met Gly Lys Glu Pro Thr Leu Gln Asn His Leu <210> 6 <211> 623 <212> PRT
<213> Homo Sapiens <220>
<221> misc feature <223> Incyte ID No: 7477587CD1 <400> 6 Met Trp Ala Pro Gly Thr Arg Gln Gln Gly Gly Pro Glu Met Ala His Tle Gln Asn Val Glu Ala His Thr Ser Ser Ala Leu Trp Gly Arg Ser Pro Arg Lys Pro Pro Thr Pro His Ala Arg Glu Ser Leu Ser Phe Pro Leu Glu Arg Pro Arg Ser Gly Arg Ser Ala Val Val Ser Ala Arg Leu Arg Gln Ser Pro Arg Met Glu Pro Arg Pro Arg Arg Arg Arg Arg Ser Arg Pro Leu Val Ala Ala Phe Leu Arg Asp Pro Gly Ser Gly Arg Va1 Tyr Arg Arg Gly Lys Leu Ile Gly Lys 95 , 100 105 Gly Ala Phe Ser Arg Cys Tyr Lys Leu Thr Asp Met Ser Thr Ser Ala Va1 Phe Ala Leu Lys Val Val Pro Cys Gly Gly Ala Gly Ala Gly Trp Leu Arg Pro Gln Gly Lys Val Glu Arg Glu Ile Ala Leu His Ser Arg Leu Arg Pro Arg Asn Ile Val Ala Phe His Gly His Phe Ala Asp Arg Asp His Val Tyr Met Val Leu G1u Tyr Cys Ser Arg Gln Ser Leu Ala His Val Leu Arg Ala Arg Gln Ile Leu Thr Glu Pro Glu Val Arg Asp Tyr Leu Arg Gly Leu Val Ser Gly Leu Arg Tyr Leu His Gln Arg Cys Ile Leu His Arg Asp Leu Lys Leu Ser Asn Phe Phe Leu Asn Lys Asn Met Glu Val Lys Ile Gly Asp Leu Gly Leu Ala Ala Lys Val Gly Pro Gly Gly Arg Cys His Arg Tyr Thr Val Leu Thr Gly Thr Pro Pro Phe Met Ala Ser Pro Leu Ser Glu Met Tyr Gln Asn Ile Arg Glu Gly His Tyr Pro Glu Pro Ala His Leu Ser Ala Asn Ala Arg Arg Leu Ile Va1 His Leu Leu Ala Pro Asn Pro Ala G1u Arg Pro Ser Leu Asp His Leu Leu Gln Asp Asp Phe Phe Thr Gln Gly Phe Thr Pro Asp Arg Leu Pro Ala His Ser Cys His Ser Pro Pro Ile Phe Ala Ile Pro.Pro Pro Leu Gly Arg Ile Phe Arg Lys Val Gly Gln Arg Leu Leu Thr Gln Cys Arg Pro Pro Cys Pro Phe Thr Pro Lys Glu Ala Ser Gly Pro Gly Glu Gly Gly Pro Asp Pro Asp Ser Met Glu Trp Asp Gly G1w Ser Ser Leu Ser Ala Lys Glu Val Pro Cys Leu Glu Gly Pro Ile His Leu Val Ala Gln G1y Thr Leu Gln Ser Asp Leu Ala Ala Thr Gln Asp Pro Leu Gly Glu Gln Gln Pro Ile Leu Trp Ala Pro Lys Trp Val Asp Tyr Ser Ser Lys Tyr Gly Phe Gly Tyr Gln Leu Leu Asp Gly Gly Arg Thr Gly Arg His Pro His Gly Pro Ala Thr Pro Arg Arg Tyr Leu Leu Ser Thr Tyr Cys Ala His Leu Gln Val Leu Pro Ala Cys Gln Val Cys Tyr Met Pro Asn Cys Gly Arg Leu Glu Ala Phe Ala Leu Arg Asp Va1 Pro Gly Leu Leu Gly Ala Lys Leu Ala Val Leu Gln Leu Phe Ala Gly Cys Leu Arg Arg Arg Leu Arg Glu Glu Gly Thr Leu Pro Thr Pro Val Pro Pro Ala G1y Pro Gly Leu Cys Leu Leu Arg Phe Leu Ala Ser Glu His Ala Leu Leu Leu Leu Phe Ser Asn Gly Met Val Gln Val Ser Phe Ser Gly Val Pro Ala Gln Leu Val Leu Ser Gly Glu Gly Glu Gly Leu Gln Leu Thr Leu Trp Glu Gln Gly Ser Pro Gly Thr Ser Tyr Ser Leu Asp Val Pro Arg Ser His Gly Cys Ala Pro Thr Thr Gly Gln His Leu His His Ala Leu Arg Met Leu Gln Ser Ile <210> 7 <211> 797 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 7594537CD1 <400> 7 Met Thr Asn Gln Glu Lys Trp Ala His Leu Ser Pro Ser Glu Phe Ser Gln Leu Gln Lys Tyr Ala Glu Tyr Ser Thr Lys Lys Leu Lys Asp Val Leu Glu Glu Phe His Gly Asn Gly Val Leu A1a Lys Tyr Asn Pro Glu Gly Thr Ile Asp Phe Glu Gly Phe Lys Leu Phe Met Lys Thr Phe Leu Glu Ala Glu Leu Pro Asp Asp Phe Thr Ala His Leu Phe Met Ser Phe Ser Asn Lys Phe Pro His Ser Ser Pro Met Val Lys Ser Lys Pro Ala Leu Leu Ser G1y Gly Leu Arg Met Asn Lys Gly Ala Ile Thr Pro Pro Arg Thr Thr Ser Pro A1a Asn Thr Cys Ser Pro Glu Val Ile His Leu Lys Asp Ile Va1 Cys Tyr Leu Ser Leu Leu Glu Arg Gly Arg Pro Glu Asp Lys Leu Glu Phe Met Phe Arg Leu Tyr Asp Thr Asp Gly Asn Gly Phe Leu Asp Ser Ser Glu Leu Glu Asn I1e Ile Ser Gln Met Met His Val Ala Glu Tyr Leu G1u Trp Asp Val Thr Glu Leu Asn Pro Ile Leu His Glu Met Met Glu Glu Ile Asp Tyr Asp His Asp Gly Thr Val Ser Leu Glu Glu Trp Ile Gln Gly Gly-Met Thr Thr Ile Pro Leu Leu Val Leu Leu Gly Leu Glu Asn Asn Val Lys Asp Asp Gly Gln His Val Trp Arg Leu Lys His Phe Asn Lys Pro Ala Tyr Cys Asn Leu Cys Leu Asn Met Leu Ile G1y Val Gly Lys Gln Gly Leu Cys Cys Ser Phe Cys Lys Tyr Thr Val His Glu Arg Cys Val Ala Arg Ala Pro Pro Ser Cys Ile Lys Thr Tyr Val Lys Ser Lys Arg Asn Thr Asp Val Met His His Tyr Trp Val Glu G1y Asn Cys Pro Thr Lys Cys Asp Lys Cys His Lys Thr Val Lys Cys Tyr Gln Gly Leu Thr G1y Leu His Cys Val Trp Cys Gln Ile Thr Leu His Asn Lys Cys Ala Ser His Leu Lys Pro Glu Cys Asp Cys Gly Pro Leu Lys Asp His Ile Leu Pro Pro Thr Thr Ile Cys Pro Val Val Leu Gln Thr Leu Pro Thr Ser Gly Val Ser Val Pro Glu Glu Arg G1n Ser Thr Val Lys Lys Glu Lys Ser Gly Ser Gln Gln Pro Asn Lys Val Ile Asp Lys Asn Lys Met Gln Arg Ala Asn Ser Val Thr Val Asp Gly Gln Gly Leu Gln Va1 Thr Pro Val Pro Gly Thr His Pro Leu Leu Val Phe Va1 Asn Pro Lys Ser Gly Gly Lys Gln Gly Glu Arg Ile Tyr Arg Lys Phe Gln Tyr Leu Leu Asn Pro Arg Gln Val Tyr Ser Leu Ser Gly Asn Gly Pro Met Pro Gly Leu Asn Phe Phe Arg Asp Val Pro Asp Phe Arg Val Leu Ala Cys Gly Gly Asp Gly Thr Val Gly Trp Val Leu Asp Cys Ile Glu Lys Ala Asn Val Gly Lys His Pro Pro Val Ala Ile Leu Pro Leu Gly Thr Gly Asn Asp Leu Ala Arg Cys Leu Arg Trp Gly Gly Gly Tyr Glu Gly Glu Asn Leu Met Lys Ile Leu Lys Asp Ile Glu Asn Ser Thr Glu Ile Met Leu Asp Arg Trp Lys Phe Glu Val Ile Pro Asn Asp Lys Asp Glu Lys Gly Asp Pro Val Pro Tyr Ser Ile Ile Asn Asn Tyr Phe Ser Ile Gly Val Asp Ala Ser=Ile Ala His Arg Phe His Ile Met Arg Glu Lys His Pro Glu Lys Phe Asn Ser Arg Met Lys Asn Lys Phe Trp Tyr Phe Glu Phe Gly Thr Ser Glu Thr Phe Ser Ala Thr Cys Lys Lys Leu His Glu Ser Val Glu Ile Glu Cys Asp Gly Val Gln Ile Asp Leu Ile Asn Ile Ser Leu Glu Gly Ile Ala Ile Leu Asn Ile Pro Ser Met His Gly Gly Ser Asn Leu Trp Gly Glu Ser Lys Lys Arg Arg Ser His Arg Arg Ile Glu Lys Lys Gly Ser Asp Lys Arg Thr Thr Val Thr Asp Ala Lys Glu Leu Lys Phe Ala Ser Gln Asp Leu Ser Asp G1n Leu Leu Glu Val Val Gly Leu Glu Gly Ala Met Glu Met Gly Gln Ile Tyr Thr Gly Leu Lys Ser Ala Gly Arg Arg Leu Ala Gln Cys Ser Cys Val Val Ile Arg Thr Ser Lys Ser Leu Pro Met Gln Ile Asp Gly Glu Pro Trp Met Gln Thr Pro Cys Thr Ile Lys Ile Thr His Lys Asn Gln Ala Pro Met Leu Met G1y Pro Pro Pro Lys Thr Gly Leu Phe Cys Ser Leu Va1 Lys Arg Thr Arg Asn Arg Ser Lys Glu <210> 8 <211> 749 <212> PRT

<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 70467491CD1 <400> 8 Met Ser Thr Arg Thr Pro Leu Pro Thr Val Asn Glu Arg Asp Thr Glu Asn Ala Val Leu Pro His Thr Ser His Gly Asp Gly Arg Gln Glu Val Thr Ser Arg Thr Ser Arg Ser Gly Ala Arg Cys Arg Asn Ser Ile Ala Ser Cys Ala Asp Glu Gln Pro His Ile Gly Asn Tyr Arg Leu Leu Lys Thr Ile Gly Lys Gly Asn Phe Ala Lys Va1 Lys Leu Ala Arg His Ile Leu Thr Gly Arg Glu Lys Asn Val Arg Ile Ser Lys Glu Ile Asp Asn Phe Leu Gly Lys His Asp Leu Pro Lys Leu Thr Leu Glu Lys Asn Arg Tyr Thr Ser Val Thr Thr G1u Val Glu Lys Val Val Asn Ile Leu Pro Asn Leu Glu Phe Met I1e Glu Phe Phe Glu Ile Tyr Ser Ile Gly Glu Val Phe Asp Tyr Leu Va1 Ala His Gly Arg Met Lys Glu Lys Glu A1a Arg Ser Lys Phe Arg Gln Ile Val Ser Ala Val Gln Tyr Cys His Gln Lys Arg Ile Val His Arg Asp Leu Lys Ala Glu Asn Leu Leu Leu Asp Ala Asp Met Asn Ile Lys Ile Ala Asp Phe Gly Phe Ser Asn Glu Phe Thr Val Gly Gly Lys Leu Asp Thr Phe Cys Gly Ser Pro Pro Tyr Ala Ala Pro Glu Leu Phe Gln Gly Lys Lys Tyr Asp Gly Pro Glu Val Asp Va1 Trp Ser Leu Gly Val Ile Leu Tyr Thr Leu Val Ser Gly Ser Leu Pro Phe Asp Gly Gln Asn Leu Lys Glu Leu Arg Glu Arg Val Leu Arg Gly Lys Tyr Arg 21e Pro Phe Tyr Met Ser Thr Asp Cys Glu Asn Leu Leu Lys Arg Phe Leu Val Leu Asn Pro Ile Lys Arg Gly Thr Leu Glu Gln Ile Met Lys Asp Arg Trp I1e Asn A1a Gly His Glu Glu Asp Glu Leu Lys Pro Phe Val Glu Pro Glu Leu Asp Ile Ser Asp Gln Lys Arg Ile Asp Ile Met Val Gly Met Gly Tyr Ser Gln Glu Glu Ile Gln Glu Ser Leu Ser Lys Met Lys Tyr Asp G1u Ile Thr Ala Thr Tyr Leu Leu Leu Gly Arg Lys Ser Ser Glu Leu Asp Ala Ser Asp Ser Ser Ser Ser Ser Asn Leu Ser Leu Ala Lys Val Arg Pro Ser Ser Asp Leu Asn Asn Ser Thr Gly Gln Ser Pro His His Lys Val Gln Arg Ser Val Ser Ser Ser Gln Lys Gln Arg Arg Tyr Ser Asp His Ala Gly Pro Ala Ile Pro Ser Val Val Ala Tyr Pro Lys Arg Ser Gln Thr Ser Thr A1a Asp~Ser Asp Leu Lys Glu Asp Gly Ile Ser Ser Arg Lys Ser Ser Gly Ser Ala Val G1y Gly Lys Gly Ile Ala Pro Ala Ser Pro Met Leu G1y Asn Ala Ser Asn Pro Asn Lys A1a Asp Ile Pro Glu Arg Lys Lys Ser Ser Thr Val Pro Ser Ser Asn Thr Ala Ser Gly Gly Met Thr Arg Arg Asn Thr Tyr Val Cys Ser Glu Arg Thr Thr Ala Asp Arg His Ser Val Ile Gln Asn Gly Lys Glu Asn Ser Thr Ile Pro Asp Gln Arg Thr Pro Val Ala Ser Thr His Ser Ile Ser Ser Ala Ala Thr Pro Asp Arg Ile Arg Phe Pro Arg Gly Thr Ala Ser Arg Ser Thr Phe His Gly Gln Pro Arg Glu Arg Arg Thr Ala Thr Tyr Asn Gly Pro Pro Ala Ser Pro Ser Leu Ser His Glu Ala Thr Pro Leu Ser Gln Thr Arg Ser Arg Gly Ser Thr Asn Leu Phe Ser Lys Leu Thr Ser Lys Leu Thr Arg Arg Leu Pro Thr Glu Tyr Glu Arg Asn Gly Arg Tyr Glu G1y Ser Ser Arg Asn Va1 Ser Ala Glu Gln Lys Asp Glu Asn Lys Glu Ala Lys Pro Arg Ser Leu Arg Phe Thr Trp Ser Met Lys Thr Thr Ser Ser Met Asp Pro Gly Asp Met Met Arg Glu Ile Arg Lys Val Leu Asp Ala Asn Asn Cys Asp Tyr Glu Gln Arg Glu Arg Phe Leu Leu Phe Cys Val His Gly Asp Gly His Ala Glu Asn Leu Val Gln Trp Glu Met Glu Val Cys Lys Leu Pro Arg Leu Ser Leu Asn Gly Val Arg Phe Lys Arg Ile Ser Gly Thr Ser Ile Ala Phe Lys Asn Ile Ala Ser Lys Tle Ala Asn Glu Leu Lys Leu <210> 9 <211> 386 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Tncyte ID No: 7478559CD1 <400> 9 Met Ala Val Pro Pro Ser Ala Pro G1n Pro Arg Ala Ser Phe His Leu Arg Arg His Thr Pro Cys Pro G1n Cys Ser Trp Gly Met Glu Glu Lys Ala Ala Ala Ser Ala Ser Cys Arg Glu Pro Pro Gly Pro Pro Arg Ala Ala Ala Val Ala Tyr Phe Gly Ile Ser Val Asp Pro Asp Asp Ile Leu Pro Gly Ala Leu Arg Leu Tle Gln Glu Leu Arg Pro His Trp Lys Pro Glu Gln Val Arg Thr Lys Arg Phe Met Asp Gly Ile Thr Asn Lys Leu Val A1a Cys Tyr Val Glu Glu Asp Met Gln Asp Cys Val Leu Val Arg Val Tyr Gly Glu Arg Thr Glu Leu Leu Val Asp Arg Glu Asn Glu Va1 Arg Asn Phe Gln Leu Leu Arg Ala His Ser Cys Ala Pro Lys Leu Tyr Cys Thr Phe Gln Asn Gly Leu Cys Tyr G1u Tyr Met Gln Gly Val Ala Leu Glu Pro Glu His Ile Arg Glu Pro Arg Leu Phe Arg Leu Ile Ala Leu Glu Met Ala Lys Ile His Thr Ile His Ala Asn Gly Ser Leu Pro Lys Pro Ile Leu Trp His Lys Met His Asn Tyr Phe Thr Leu Val Lys Asn Glu Ile Asn Pro Ser Leu Ser Ala Asp Val Pro Lys Val Glu Val Leu Glu Arg Glu Leu Ala Trp Leu Lys Glu His Leu Ser Gln Leu Glu Ser Pro Val Val Phe Cys His Asn Asp Leu Leu Cys Lys Asn Ile Ile Tyr Asp Ser Ile Lys Gly His Val Arg Phe Ile Asp Tyr Glu Tyr Ala Gly Tyr Asn Tyr Gln Ala Phe Asp Ile Gly Asn His Phe Asn Glu Phe Ala Gly Val Asn Glu Va1 Asp Tyr Cys Leu Tyr Pro Ala Arg Glu Thr Gln Leu Gln Trp Leu His Tyr Tyr Leu Gln Ala Gln Lys Gly Met Ala Val Thr Pro Arg Glu Val Gln Arg Leu Tyr Val Gln Val Asn Lys Phe Ala Leu Ala Ser His Phe Phe Trp A1a Leu Trp Ala Leu Ile Gln Asn Gln Tyr Ser Thr Ile Asp Phe Asp Phe Leu Arg Tyr Ala Val Ile Arg Phe Asn Gln Tyr Phe Lys Val Lys Pro Gln Ala Ser Ala Leu Glu Met Pro Lys <210> 10 <211> 342 <212> PRT
<213> Homo Sapiens <220>
<221> misc feature <223> Incyte ID No: 1698381CD1 <400> 10 Met Glu Lys Tyr Glu Lys Leu Ala Lys Thr Gly Glu Gly Ser Tyr Gly Val Val Phe Lys Cys Arg Asn Lys Thr Ser Gly Gln Val Val Ala Val Lys Lys Phe Val Glu Ser G1u Asp Asp Pro Val Val Lys Lys Ile Ala Leu Arg Glu Ile Arg Met Leu Lys Gln Leu Lys His Pro Asn Leu Val Asn Leu Ile Glu Val Phe Arg Arg Lys Arg Lys Met His Leu Val Phe Glu Tyr Cys Asp His Thr Leu Leu Asn Glu Leu Glu Arg Asn Pro Asn Gly Val Ala Asp Gly Val Ile Lys Ser Va1 Leu Trp Gln Thr Leu Gln Ala Leu Asn Phe Cys His Ile His Asn Cys 21e His Arg Asp Ile Lys Pro G1u Asn Ile Leu Ile Thr Lys Gln Gly Ile I1e Lys Ile Cys Asp Phe Gly Phe Ala Gln Ile Leu Ile Pro Gly Asp Ala Tyr Thr Asp Tyr Va1 Ala Thr Arg Trp Tyr Arg Ala Pro Glu Leu Leu Val Gly Asp Thr Gln Tyr Gly Ser Ser Val Asp Ile ,Trp Ala Ile Gly Cys Val Phe Ala Glu Leu Leu Thr Gly Gln Pro Leu Trp Pro Gly Lys Ser Asp Val Asp Gln Leu Tyr Leu Ile Ile Arg Thr Leu Gly Lys Leu Ile Pro Arg His Gln Ser I1e Phe Lys Ser Asn Gly Phe Phe His Gly Ile Ser Ile Pro Glu Pro Glu Asp Met Glu Thr Leu Glu Glu Lys Phe Ser Asp Val His Pro Val A1a Leu Asn Phe Met Lys Gly Cys Leu Lys Met Asn Pro Asp Asp Arg Leu Thr Cys Ser Gln Leu Leu Glu Ser Ser Tyr Phe Asp Ser Phe Gln Glu Ala Gln I1e Lys Arg Lys Ala Arg Asn 290 295 ~ 300 Glu Gly Arg Asn Arg Arg Arg Gln Gln Asn Gln Leu Leu Pro Leu Ile Pro Gly Ser His Ile Ser Pro Thr Pro Asp Gly Arg Lys Gln Val Leu Gln Leu Lys Phe Asp His Leu Pro Asn Ile <210> 11 <211> 1164 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 7474637CD1 <400> 11 , Met Ala Gly Ala Gly Gly Gln His His Pro Pro G1y Ala Ala Gly Gly Ala A1a Ala Gly Ala Gly Ala Ala Va1 Thr Ser Ala Ala Ala Ser Ala Gly Pro Gly Glu Asp Ser Ser Asp Ser Glu Ala Glu Gln Glu Gly Pro Gln Lys Leu Ile Arg Lys Val Ser Thr Ser Gly Gln Ile Arg Thr Lys Thr Ser Ile Lys Glu Gly Gln Leu Leu Lys Gln Thr Ser Ser Phe Gln Arg Trp Lys Lys Arg Tyr Phe Lys Leu Arg Gly Arg Thr Leu Tyr Tyr Ala Lys Asp Ser Lys Ser Leu Ile Phe 95 100 ~ 105 Asp Glu Val Asp Leu Ser Asp Ala Ser Val Ala Glu Ala Ser Thr Lys Asn A1a Asn Asn Ser Phe Thr Ile Ile Thr Pro Phe Arg Arg Leu Met Leu Cys Ala Glu Asn Arg Lys Glu Met Glu Asp Trp Ile Ser Ser Leu Lys Ser Va1 Gln Thr Arg Glu Pro Tyr Glu Val Ala Gln Phe Asn Val Glu=His Phe Ser Gly Met His Asn Trp Tyr Ala Cys Ser His Ala Arg Pro Thr Phe Cys Asn Val Cys Arg Glu Ser Leu Ser Gly Val Thr Ser His Gly Leu Ser Cys Glu Val Cys Lys Phe Lys A1a His Lys Arg Cys A1a Val Arg Ala Thr Asn Asn Cys Lys Trp Thr Thr Leu Ala Ser I1e Gly Lys Asp Ile Ile Glu Asp Glu Asp Gly Val Ala Met Pro His G1n Trp Leu Glu Gly Asn Leu Pro Val Ser Ala Lys Cys Ala Val Cys Asp Lys Thr Cys Gly Ser Val Leu Arg Leu Gln Asp Trp Lys Cys Leu Trp Cys Lys Thr Met Val His Thr Ala Cys Lys Asp Leu Tyr His Pro Ile Cys Pro Leu Gly Gln Cys Lys Val Ser Ile Ile Pro Pro Ile Ala Leu Asn Ser Thr Asp Ser Asp Gly Phe Cys Arg Ala Thr Phe Ser Phe Cys Val Ser Pro Leu Leu Val Phe Val Asn Ser Lys Ser Gly Asp Asn Gln Gly Val Lys Phe Leu Arg Arg Phe Lys Gln Leu Leu Asn Pro Ala Gln Val Phe Asp Leu Met Asn Gly Gly Pro His Leu Gly Leu Arg Leu Phe Gln Lys Phe Asp Asn Phe Arg Ile Leu Val Cys Gly Gly Asp Gly Ser Val Gly Trp Val Leu Ser Glu Ile Asp Lys Leu Asn Leu Asn Lys Gln Cys Gln Leu Gly Val Leu Pro Leu Gly Thr G1y Asn Asp Leu Ala Arg Val Leu Gly Trp Gly Gly Ser Tyr Asp Asp Asp Thr Gln Leu Pro Gln Ile Leu Glu Lys Leu Glu Arg Ala Ser Thr Lys Met Leu Asp Arg Trp Ser Ile Met Thr Tyr Glu Leu Lys Leu Pro Pro Lys Ala Ser Leu Leu Pro Gly Pro Pro Glu Ala Ser Glu Glu Phe Tyr Met Thr I1e Tyr Glu Asp Ser Val Ala Thr His Leu Thr Lys I1e Leu Asn Ser Asp Glu His Ala Val Val Ile Ser Ser A1a Lys Thr Leu Cys Glu Thr Val Lys Asp Phe Val Ala Lys Val Glu Lys Thr Tyr Asp Lys Thr Leu Glu Asn Ala Val Val Ala Asp Ala Val Ala Ser Lys Cys Ser Val Leu Asn Glu Lys Leu G1u Gln Leu Leu Gln Ala Leu His Thr Asp Ser Gln Ala Ala Pro Val Leu Pro Gly Leu Ser Pro Leu Ile Va1 Glu Glu Asp Ala Val G1u Ser Ser Ser G1u Glu Ser Leu Gly Glu Ser Lys Glu Gln Leu Gly Asp Asp Val Thr Lys Pro Ser Ser Gln Lys Ala Val Lys Pro Arg Glu Ile Met Leu Arg Ala Asn Ser Leu Lys Lys Ala Val Arg G1n Val Ile Glu Glu Ala Gly Lys Val Met Asp Asp Pro Thr Val His Pro Cys Glu Pro Ala Asn Gln Ser Ser Asp Tyr Asp Ser Thr Glu Thr Asp Glu Ser Lys Glu Glu Ala Lys Asp Asp Gly Ala Lys Glu Ser I1e Thr Val Lys Thr Ala Pro Arg Ser Pro Asp Ala Arg Ala Ser Tyr Gly His Ser Gln Thr Asp Ser Val Pro Gly Pro Ala Val Ala Ala Ser Lys Glu Asn Leu Pro Val Leu Asn Thr Arg Ile Ile Cys Pro Gly Leu Arg Ala Gly Leu Ala Ala Ser Ile Ala Gly Ser Ser Ile Ile Asn Lys Met Leu Leu Ala Asn Ile Asp Pro Phe Gly Ala Thr Pro Phe Ile Asp Pro Asp Leu Asp Ser Val Asp Gly Tyr Ser Glu Lys Cys Val Met Asn Asn Tyr Phe Gly Ile Gly Leu Asp Ala Lys Ile Ser Leu Glu Phe Asn Asn Lys Arg Glu Glu His Pro Glu Lys Cys Arg Ser Arg Thr Lys Asn Leu Met Trp Tyr Gly Va1 Leu Gly Thr Arg Glu Leu Leu Gln Arg Ser Tyr Lys Asn Leu Glu Gln Arg Val Gln Leu Glu Cys Asp Gly Gln Tyr Ile Pro Leu Pro Ser Leu Gln Gly Ile Ala Val Leu Asn Ile Pro Ser Tyr Ala Gly Gly Thr Asn Phe Trp Gly Gly Thr Lys Glu Asp Asp Ile Phe Ala Ala Pro Ser Phe Asp Asp Lys Ile Leu Glu Va1 Val Ala Ile Phe Asp Ser Met Gln Met Ala Val Ser Arg Val Ile Lys Leu Gln His His Arg Ile Ala Gln Cys Arg Thr Val Lys I1e Thr Ile Phe Gly 905 , 910 915 Asp Glu Gly Val Pro Val Gln Val Asp Gly Glu Ala Trp Val Gln Pro Pro Gly Ile Ile Lys Ile Val His Lys Asn Arg Ala Gln Met Leu Thr Arg Asp Arg Ala Phe Glu Ser Thr Leu Lys Ser Trp Glu Asp Lys Gln Lys Cys Asp Ser G1y Lys Pro Va1 Leu Arg Thr His Leu Tyr Ile His His Ala Ile Asp Leu Ala Thr Glu Glu Va1 Ser Gln Met Gln Leu Cys Ser Gln Ala Ala Glu Glu Leu Ile Thr Arg Ile Cys Asp Ala Ala Thr Ile His Cys Leu Leu Glu Gln Glu Leu Ala His Ala Val Asn Ala Cys Ser His Ala Leu Asn Lys Ala Asn Pro Arg Cys Pro Glu Ser Leu Thr Arg Asp Thr Ala Thr Glu Ile A1a Ile Asn Va1 Lys Ala Leu Tyr Asn Glu Thr Glu Ser Leu Leu Val Gly Arg Va1 Pro Leu Gln Leu Glu Ser Pro His Glu Glu Arg Val Ser Asn Ala Leu His Ser Val Glu Val Glu Leu Gln Lys Leu Thr Glu Ile Pro Trp Leu Tyr Tyr Ile Leu His Pro Asn Glu Asp Glu Glu Pro Pro Met Asp Cys Thr Lys Arg Asn Asn Arg Ser Thr Val Phe Arg Ile Val Pro Lys Phe Lys Lys Glu Lys Val Gln Lys Gln Lys Thr Ser Ser Gln Pro Gly Ser Gly Asp Thr G1u Ser Gly Ser Cys Glu Ala Asn Ser Pro Gly Asn <210> 12 <211> 268 <212> PRT
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 7170260CD1 <400> 12 Met Glu Asp Phe Leu Leu Ser Asn Gly Tyr Gln Leu Gly Lys Thr Ile Gly Glu Gly Thr Tyr Ser Lys Val Lys Glu Ala Phe Ser Lys Lys His Gln Arg Lys Val Ala Ile Lys Val Ile Asp Lys Met Gly Gly Pro Glu Glu Phe Ile Gln Arg Phe Leu Pro Arg Glu Leu Gln Ile Val Arg Thr Leu Asp His Lys Asn Ile Ile Gln Val Tyr Glu Met Leu Glu Ser Ala Asp Gly Lys Ile Cys Leu Val Met Glu Leu Ala Glu Gly Gly Asp Val Phe Asp Cys Val Leu Asn Gly Gly Pro Leu Pro Glu Ser Arg Ala Lys Ala Leu Phe Arg Gln Met Val Glu Ala I1e Arg Tyr Cys His Gly Cys G1y Val Ala His Arg Asp Leu Lys Cys Glu Asn A1a Leu Leu Gln Gly Phe Asn Leu Lys Leu Thr Asp Phe Gly Phe Ala Lys Val Leu Pro Lys Ser His Arg Glu Leu Ser Gln Thr Phe Cys Gly Ser Thr Ala Tyr Ala Ala Pro Glu Val Leu Gln Gly Ile Pro His Asp Ser Lys Lys Gly Asp Val Trp Ser Met Gly Val Val Leu Tyr Val Met Leu Cys Ala Ser Leu Pro Phe Asp Asp Thr Asp Ile Pro Lys Met Leu Trp Gln Gln Gln Lys Gly Val Ser Phe Pro Thr His Leu Ser Ile Ser Ala Asp Cys Gln Asp Leu Leu Lys Arg Leu Leu Glu Pro Asp Met Ile Leu Arg Pro Ser Ile Glu Glu Val Ser Trp His Pro Trp Leu Ala Ser Thr <210> 13 <211> 965 <212> PRT
<213> Homo Sapiens <220>
<221> misc feature <223> Incyte ID No: 1797506CD1 <400> 13 Met Arg Arg Ala Gly Ile Gly Glu Asp Ser Arg Leu G1y Leu Gln Ala Gln Pro Gly Ala Glu Pro Ser Pro Gly Arg Ala Gly Thr Glu Arg Ser Leu Gly Gly Thr Gln Gly Pro Gly Gln Pro Cys Ser Cys Pro Gly Ala Met Ala Ser Ala Val Arg Gly Ser Arg Pro Trp Pro Arg Leu Gly Leu Gln Leu G1n Phe Ala Ala Leu Leu Leu Gly Thr Leu Ser Pro Gln Val His Thr Leu Arg Pro Glu Asn Leu Leu Leu Va1 Ser Thr Leu Asp Gly Ser Leu His Ala Leu Ser Lys Gln Thr Gly Asp Leu Lys Trp Thr Leu Arg Asp Asp Pro Val I1e Glu Gly Pro Met Tyr Val Thr Glu Met Ala Phe Leu Ser Asp Pro Ala Asp Gly Ser Leu Tyr Ile Leu Gly Thr Gln Lys Gln Gln Gly Leu Met Lys Leu Pro Phe Thr Ile Pro Glu Leu Val His Ala Ser Pro Cys Arg Ser Ser Asp Gly Val Phe Tyr Thr Gly Arg Lys Gln Asp Ala Trp Phe Val Val Asp Pro Glu Ser Gly Glu Thr Gln Met Thr Leu Thr Thr G1u Gly Pro Ser Thr Pro Arg Leu Tyr Ile G1y Arg Thr Gln Tyr Thr Val Thr Met His Asp Pro Arg Ala Pro Ala Leu Arg Trp Asn Thr Thr Tyr Arg Arg Tyr Ser Ala Pro Pro Met Asp Gly Ser Pro G1y Lys Tyr Met Ser His Leu Ala Ser Cys G1y Met Gly Leu Leu Leu Thr Val Asp Pro Gly Ser Gly Thr Val Leu Trp Thr Gln Asp Leu Gly Val Pro Va1 Met Gly Val Tyr Thr Trp His Gln Asp G1y Leu Arg Gln Leu Pro His Leu Thr Leu Ala Arg Asp Thr Leu His Phe Leu Ala Leu Arg Trp Gly His Ile Arg Leu Pro Ala Ser Gly Pro Arg Asp Thr Ala Thr Leu Phe Ser Thr Leu Asp Thr 320 325 ' 330 Gln Leu Leu Met Thr Leu Tyr Val Gly Lys Asp Glu Thr Gly Phe Tyr Val Ser Lys Ala Leu Val His Thr Gly Val Ala Leu Val Pro Arg Gly Leu Thr Leu Ala Pro Ala Asp Gly Pro Thr Thr Asp Glu Val Thr Leu Gln Val Ser Gly G1u Arg Glu Gly Ser Pro Ser Thr Ala Val Arg Tyr Pro Ser Gly Ser Val Ala Leu Pro Ser Gln Trp Leu Leu Ile Gly His His Glu Leu Pro Pro Val Leu His Thr Thr Met Leu Arg Val His Pro Thr Leu Gly Ser Gly Thr Ala Glu Thr Arg Pro Pro Glu Asn Thr Gln Ala Pro Ala Phe Phe Leu Glu Leu Leu Ser Leu Ser Arg Glu Lys Leu Trp Asp Ser Glu Leu His Pro Glu Glu Lys Thr Pro Asp Ser Tyr Leu Gly Leu Gly Pro Gln Asp Leu Leu Ala Ala Ser Leu Thr Ala Val Leu Leu Gly Gly Trp Ile Leu Phe Val Met Arg Gln Gln Gln Glu Thr Pro Leu Ala Pro Ala Asp Phe Ala His I1e Ser Gln Asp Ala Gln Ser Leu His Ser Gly Ala Ser Arg Arg Ser Gln Lys Arg Leu Gln Ser Pro Ser Pro Glu Ser Pro Pro Ser Ser Pro Pro Ala Glu Gln Leu Thr Val Val Gly Lys Ile Ser Phe Asn Pro Lys Asp Val Leu Gly Arg Gly Ala Gly Gly Thr Phe Va1 Phe Arg Gly Gln Phe Glu Gly Arg Ala Val A1a Val Lys Arg Leu Leu Arg Glu Cys Phe Gly Leu Val Arg Arg Glu Val Gln Leu Leu Gln Glu Ser Asp Arg His Pro Asn Val Leu Arg Tyr Phe Cys Thr Glu Arg Gly Pro Gln Phe His Tyr Ile Ala Leu Glu Leu Cys Arg Ala Ser Leu Gln Glu Tyr Val Glu Asn Pro Asp Leu Asp Arg Gly Gly Leu Glu Pro Glu Val Val Leu Gln Gln Leu Met Ser G1y Leu Ala His Leu His Ser Leu His Ile Val His Arg Asp Leu Lys Pro Gly Asn Ile Leu Ile Thr Gly Pro Asp Ser G1n Gly Leu Gly Arg Val Val Leu Ser Asp Phe Gly Leu Cys Lys Lys Leu Pro Ala Gly Arg Cys Ser Phe Ser Leu His Ser Gly Ile Pro Gly Thr Glu Gly Trp Met A1a Pro Glu Leu Leu Gln Leu Leu Pro Pro Asp Ser Pro Thr Ser Ala Va1 Asp Ile Phe Ser Ala Gly Cys Val Phe Tyr Tyr Val Leu Ser Gly GIy Ser His Pro Phe Gly Asp Ser Leu Tyr Arg Gln Ala Asn Ile Leu Thr G1y Ala Pro Cys Leu Ala His Leu Glu Glu G1u Val His Asp Lys Val Val Ala Arg Asp Leu Val Gly Ala Met Leu Ser Pro Leu Pro Gln Pro Arg Pro Ser Ala Pro Gln Val Leu Ala His Pro Phe Phe Trp Ser Arg Ala Lys Gln Leu Gln Phe Phe Gln Asp Val Ser Asp Trp Leu Glu Lys Glu Ser Glu Gln Glu Pro Leu Val Arg Ala Leu Glu Ala Gly Gly Cys Ala Val Val Arg Asp Asn Trp His Glu His Ile Ser Met Pro Leu Gln Thr Asp Leu Arg Lys'Phe Arg Ser Tyr Lys Gly Thr Ser Val Arg Asp Leu Leu Arg Ala Val Arg Asn Lys Lys His His Tyr Arg Glu Leu Pro Val Glu Val Arg Gln Ala Leu Gly Gln Val Pro Asp Gly Phe Val Gln Tyr Phe Thr Asn Arg Phe Pro Arg Leu Leu Leu His Thr His Arg Ala Met Arg Ser Cys Ala Ser Glu Ser Leu Phe Leu Pro Tyr Tyr Pro Pro Asp Ser Glu A1a Arg Arg Pro Cys Pro Gly Ala Thr Gly Arg <210> 14 <211> 329 <212> PRT
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 1851973CD1 <400> l4 Met Asp Pro Thr Ala Gly Ser Lys Lys Glu Pro Gly Gly Gly Ala Ala Thr Glu Glu Gly Val Asn Arg Ile Ala Val Pro Lys Pro Pro Ser Ile Glu Glu Phe Ser Ile Val Lys Pro Ile Ser Arg Gly Ala Phe Gly Lys Val Tyr Leu Gly Gln Lys Gly Gly Lys Leu Tyr Ala Val Lys Val Val Lys Lys Ala Asp Met Ile Asn Lys Asn Met Thr His G1n Val Gln Ala Glu Arg Asp A1a Leu Ala Leu Sex Lys Ser Pro Phe Ile Val His Leu Tyr Tyr Ser Leu Gln Ser Ala Asn Asn Val Tyr Leu Val Met Glu Tyr Leu Ile Gly Gly Asp Val Lys Ser Leu Leu His Ile Tyr Gly Tyr Phe Asp Glu Glu Met Ala Val Lys Tyr Ile Ser Glu Val Ala Leu Ala Leu Asp Tyr Leu His Arg His Gly Ile Ile His Arg Asp Leu Lys Pro Asp Asn Met Leu Ile Ser Asn G1u Gly His Ile Lys Leu Thr Asp Phe Gly Leu Ser Lys Val Thr Leu Asn Arg Asp Ile Asn Met Met Asp Ile Leu Thr Thr Pro Ser Met A1a Lys Pro Arg Gln Asp Tyr Ser Arg Thr Pro Gly Gln Val Leu Ser Leu Ile Ser Ser Leu Gly Phe Asn Thr Pro Ile Ala Glu Lys Asn Gln Asp Pro Ala Asn I1e Leu Ser Ala Cys Leu Ser Glu Thr Ser Gln Leu Ser Gln Gly Leu Val Cys Pro Met Ser Val Asp Gln Lys Asp Thr Thr Pro Tyr Ser Ser Lys Leu Leu Lys Ser Cys Leu Glu Thr Val Ala Ser Asn Pro Gly Met Pro Val Lys Cys Leu Thr Ser Asn Leu Leu Gln Ser Arg Lys Arg Leu Ala Thr Ser Ser Ala Ser Ser Gln Ser His Thr Phe I1e Ser Ser Val Glu Ser Glu Cys His Ser Ser Pro Lys Trp Glu Lys Asp Cys Gln Val <210> 15 <211> 945 <212> PRT
<213> Homo sapiens <220>
<221> misc feature <223> Incyte ID No: 7474604CD1 <400> 15 Met Thr Lys Ser Glu Glu Gln Gln Pro Leu Ser Leu Gln Lys Ala Leu Gln Gln Cys Glu Leu Val G1n Asn Met Ile Asp Leu Ser Ile Ser Asn Leu Glu Gly Leu Arg Thr Lys Cys Ala Thr Ser Asn Asp Leu Thr Gln Lys Glu Ile Arg Thr Leu Glu Ser Lys Leu Val Lys Tyr Phe Ser Arg Gln Leu Ser Cys Lys Lys Lys Val Ala Leu Gln Glu Arg Asn Ala Glu Leu Asp Gly Phe Pro G1n Leu Arg His Trp Phe Arg Ile Val Asp Val Arg Lys Glu Val Leu Glu Glu Ile Ser Pro Gly G1n Leu Ser Leu Glu Asp Leu Leu Glu Met Thr Asp Glu G1n Val Cys Glu Thr Val Glu Lys Tyr Gly Ala Asn Arg Glu Glu Cys Ala Arg Leu Asn Ala Ser Leu Ser Cys Leu Arg Asn Val His Met Ser G1y Gly Asn Leu Ser Lys Gln Asp Trp Thr I1e Gln Trp Pro Thr Thr Glu Thr Gly Lys Glu Asn Asn Pro Val Cys Pro Pro Glu Pro Thr Pro Trp Ile Arg Thr His Leu Ser Gln Ser Pro Arg Val Pro Ser Lys Cys Va1 Gln His Tyr Cys His Thr Ser Pro Thr Pro Gly Ala Pro Val Tyr Thr His Val Asp Arg Leu Thr Va1 Asp Ala Tyr Pro Gly Leu Cys Pro Pro Pro Pro Leu Glu Ser Gly His Arg Ser Leu Pro Pro Ser Pro Arg Gln Arg His Ala Val Arg Thr Pro Pro Arg Thr Pro Asn Ile Val Thr Thr Val Thr Pro Pro Gly Thr Pro Pro Met Arg Lys Lys Asn Lys Leu Lys Pro Pro Gly Thr Pro Pro Pro Ser Ser Arg Lys Leu Ile His Leu Ile Pro Gly Phe Thr A1a Leu His Arg Ser Lys Ser His Glu Phe Gln Leu Gly His Arg Val Asp Glu Ala His Thr Pro Lys Ala Lys Lys Lys Ser Lys Pro Leu Asn Leu Lys Ile His Ser Ser Val Gly Ser Cys Glu Asn Ile Pro Ser Gln Gln Arg Ser Pro Leu Leu Ser Glu Arg Ser Leu Arg Ser Phe Phe Val Gly His Ala Pro Phe Leu Pro 5er Thr Pro Pro Val His Thr Glu Ala Asn Phe Ser Ala Asn Thr Leu Ser Val Pro Arg Trp Ser Pro G1n Ile Pro Arg Arg Asp Leu Gly Asn Ser Ile Lys His Arg Phe Ser Thr Lys Tyr Trp Met Sex G1n Thr Cys Thr Val Cys Gly Lys G1y Met Leu Phe Gly Leu Lys Cys Lys Asn Cys Lys Leu Lys Cys His Asn Lys Cys Thr Lys Glu Ala Pro Pro Cys His Leu Leu Ile Ile His Arg Gly Asp Pro Ala Arg Leu Val Arg Thr Glu Ser Val Pro Cys Asp Ile Asn Asn Pro Leu Arg Lys Pro Pro Arg Tyr Ser Asp Leu His Ile Ser Gln Thr Leu Pro Lys Thr Asn Lys Ile Asn Lys Asp His Ile Pro Va1 Pro Tyr Gln Pro Asp Ser Ser Ser Asn Pro Ser Ser Thr Thr Ser Ser Thr Pro Ser Ser Pro Ala Pro Pro Leu Pro Pro Ser A1a Thr Pro Pro Ser Pro Leu His Pro Ser Pro Gln Cys Thr Arg Gln Gln Lys Asn Phe Asn Leu Pro Ala Ser His Tyr Tyr Lys Tyr Lys Gln Gln Phe Ile Phe 5&0 5&5 570 Pro Asp Val Val Pro Val Pro G1u Thr Pro Thr Arg Ala Pro Gln Val Ile Leu His Pro Val Thr Ser Asn Pro Ile Leu Glu Gly Asn Pro Leu Leu Gln Ile Glu Val Glu Pro Thr Ser Glu Asn Glu Glu Val His Asp Glu Ala Glu Glu Ser Glu Asp Asp Phe Glu Glu Met Asn Leu Ser Leu Leu Ser Ala Arg Ser Phe Pro Arg Lys Ala Ser Gln Thr Ser Ile Phe Leu Gln Glu Trp Asp Ile Pro Phe Glu Gln Leu Glu Ile Gly Glu Leu Ile Gly Lys Gly Arg Phe Gly Gln Val Tyr His Gly Arg Trp His Gly Glu Val Ala Ile Arg Leu Ile Asp Ile Glu Arg Asp Asn Glu Asp Gln Leu Lys Ala Phe Lys Arg Glu Val Met A1a Tyr Arg Gln Thr Arg His G1u Asn Val Val Leu Phe Met Gly Ala Cys Met Ser Pro Pro His Leu Ala Ile Ile Thr Ser Leu Cys Lys Gly Arg Thr Leu Tyr Ser Val Val Arg Asp Ala Lys Ile Val Leu Asp Val Asn Lys Thr Arg Gln Ile Ala Gln Glu Ile Val Lys Gly Met Gly Tyr Leu His Ala Lys Gly Ile Leu His Lys Asp Leu Lys Ser Lys Asn Val Phe Tyr Asp Asn Gly Lys Val Val Ile Thr Asp Phe Gly Leu Phe Ser Ile Ser Gly Val Leu Gln Ala Gly Arg Arg Glu Asp Lys Leu Arg I1e Gln Asn G1y Trp Leu Cys His Leu Ala Pro Glu Ile Ile Arg Gln Leu Ser Pro Asp Thr Glu Glu Asp Lys Leu Pro Phe Ser Lys His Ser Asp Val Phe Ala Leu Gly Thr Ile Trp Tyr Glu Leu His Ala Arg Glu Trp Pro Phe Lys Thr Gln Pro Ala Glu Ala Ile Ile Trp Gln Met Gly Thr Gly Met Lys Pro Asn Leu Ser Gln Ile Gly Met Gly Lys Glu Ile Ser Asp Ile Leu Leu Phe Cys Trp Ala Phe Glu Gln Glu Glu Arg Pro Thr Phe Thr Lys Leu Met Asp Met Leu Glu Lys Leu Pro Lys Arg Asn Arg Arg Leu Ser His Pro Gly His Phe Trp Lys Ser Ala Glu Leu <210> 16 <211> 1009 <212> PRT
<213> Homo Sapiens <220>
<221> misc feature <223> Incyte ID No: 7474721CD1 <400> 16 Met Glu Thr Cys A1a Gly Pro His Pro Leu Arg Leu Phe Leu Cys Arg Met Gln Leu Cys Leu Ala Leu Leu Leu Gly Pro Trp Arg Pro Gly Thr Ala Glu Glu Val Ile Leu Leu Asp Ser Lys Ala Ser Gln Ala GIu Leu Gly Trp Thr Ala Leu Pro Ser Asn Gly Trp Glu Glu Ile Ser Gly Val Asp G1u His Asp Arg Pro I1e Arg Thr Tyr Gln Val Cys Asn Val Leu Glu Pro Asn Gln Asp Asn Trp Leu Gln Thr Gly Trp Ile Ser Arg Gly Arg Gly Gln Arg Ile Phe Val Glu Leu Gln Phe Thr Leu Arg Asp Cys Ser Ser Ile Pro G1y Ala A1a G1y Thr Cys Lys Glu Thr Phe Asn Val Tyr Tyr Leu Glu Thr Glu A1a Asp Leu Gly Arg Gly Arg Pro Arg Leu Gly Gly Ser Arg Pro Arg Lys Ile Asp Thr Ile Ala Ala Asp Glu Ser Phe Thr Gln Gly Asp Leu Gly Glu Arg Lys Met Lys Leu Asn Thr Glu Val Arg Glu Ile Gly Pro Leu Ser Arg Arg Gly Phe His Leu Ala Phe Gln Asp Val G1y Ala Cys Val Ala Leu Val Ser Val Arg Val Tyr Tyr Lys Gln Cys Arg Ala Thr Val Arg Gly Leu Ala Thr Phe Pro Ala Thr Ala Ala Glu Ser Ala Phe Ser Thr Leu Val Glu Val Ala Gly Thr Cys Val Ala His Ser Glu Gly Glu Pro Gly Ser Pro Pro Arg Met His Cys Gly Ala Asp Gly Glu Trp Leu Val Pro Val Gly Arg Cys Ser Cys Ser Ala Gly Phe Gln Glu Arg Gly Asp Ile Cys G1u Ala Cys Pro Pro Gly Phe Tyr Lys Val Ser Pro Arg Arg Arg Val Cys Ser Pro Cys Pro Glu His Ser Arg Ala Leu Glu Asn Ala Ser Thr Phe Cys Val Cys Gln Asp Ser Tyr Ala Arg Ser Pro Thr Asp Pro Pro Ser Ala Ser Cys Thr Arg Gly Pro Pro Ser Ala Pro Arg Asp Leu Gln Tyr Ser Leu Ser Arg Ser Pro Leu Val Leu Arg Leu Arg Trp Leu Pro Pro Ala Asp Ser Gly Gly Arg Ser Asp Val Thr Tyr Ser Leu Leu Cys Leu Arg Cys Gly Arg Glu Gly Pro Ala Gly Ala Cys Glu Pro Cys Gly Pro Arg Val Ala Phe Leu Pro Arg Gln Ala Gly Leu Arg Glu Arg Ala Ala Thr Leu Leu His Leu Arg Pro Gly Ala Arg Tyr Thr Val Arg Val Ala Va1 Leu Asn Gly Val Ser Gly Pro Ala Ala Ala Leu Val Pro Val Gly Ala Val Sex Ile Asn Pro Gly Thr Va1 G1y Pro Val Pro Val Ala Gly Val Ile Arg Asp Arg Val Glu Pro Gln Ser Val Ser Leu Ser Trp Arg Glu Pro Ile Pro Ala Gly Ala Pro Gly Ala Asn Asp Thr Glu Tyr Glu Ile Arg Tyr Tyr Glu Lys Val Gln Ser Glu Gln Thr Tyr Ser Met Val Lys Thr Gly Ala Pro Thr Val Thr Val Thr Asn Leu Lys Pro Ala Thr Arg Tyr Val Phe Gln Ile Arg Ala Ala Ser Pro Gly Pro Ser Trp Glu Ala Gln Ser Phe Asn Pro Ser Ile Glu Val Gln Thr Leu Gly Glu Ala Ala Ser G1y Ser Arg Asp Gln Ser Pro Ala I1e Val Val Thr Val Val Thr Ile Ser Ala Leu Leu Val Leu Gly Sex Val Met Ser Val Leu Ala Ile Trp Arg Arg Pro Cys Ser Tyr Gly Lys Gly Gly Gly Asp Ala His Asp Glu Glu Glu Leu Tyr Phe His Phe Lys Val Pro Thr Arg Arg Thr Phe Leu Asp Pro G1n Ser Cys Gly Asp Leu Leu Gln Ala Va1 His Leu Phe Ala Lys Glu Leu Asp Ala Lys Ser Val Thr Leu Glu Arg Ser Leu G1y Gly Gly Arg Phe Gly GIu Leu Cys Cys Gly Cys Leu Gln Leu Pro Gly Arg Gln Glu Leu Leu Val Ala Val His Met Leu Arg Asp Ser Ala Ser Asp Ser Gln Arg Leu Gly Phe Leu Ala Glu A1a Leu Thr Leu Gly G1n Phe Asp His Ser His Ile Val Arg Leu Glu Gly Val Val Thr Arg G1y Ser Thr Leu Met Ile Val Thr Glu Tyr Met Ser His Gly Ala Leu Asp Gly Phe Leu Arg Arg His Glu Gly Gln Leu Val Ala Gly Gln Leu Met Gly Leu Leu Pro Gly Leu Ala Ser Ala Met Lys Tyr Leu Ser Glu Met Gly Tyr Val His Arg Gly Leu Ala Ala Arg His Val Leu Val Ser Ser Asp Leu Val Cys Lys Ile Ser Gly Phe Gly Arg Gly Pro Arg Asp Arg Ser G1u Ala Val Tyr Thr Thr Met Ser Gly Arg Ser Pro Ala 800 805 ~ 810 Leu Trp Ala Ala Pro Glu Thr Leu Gln Phe Gly His Phe Ser Ser Ala Ser Asp Val Trp Ser Phe Gly Ile Ile Met Trp Glu Val Met Ala Phe Gly Glu Arg Pro Tyr Trp Asp Met Ser Gly Gln Asp Val Ile Lys Ala Val Glu Asp Gly Phe Arg Leu Pro Pro Pro Arg Asn Cys Pro Asn Leu Leu His Arg Leu Met Leu Asp Cys Trp Gln Lys Asp Pro Gly Glu Arg Pro Arg Phe Ser Gln Ile His Ser Ile Leu Ser Lys Met Val Gln Asp Pro Glu Pro Pro Lys Cys Ala Leu Thr Thr Cys Pro Arg Pro Pro Thr Pro Leu Ala Asp Arg Ala Phe Ser Thr Phe Pro Ser Phe Gly Ser Val Gly Ala Trp Leu G1u Ala Leu Asp Leu Cys Arg Tyr Lys Asp Ser Phe Ala Ala Ala Gly Tyr Gly Ser Leu Glu Ala Val Ala Glu Met Thr Ala Gln Arg Asp Leu Val Ser Leu Gly Ile Ser Leu Ala Glu His Arg Glu Ala Leu Leu Ser Gly Ile Ser Ala Leu Gln Ala Arg Val Leu Gln Leu Gln Gly Gln Gly Val Gln Val <220> 17 <211> 917 <212> PRT
<213> Homo Sapiens <220>
<222> misc_feature <223> Incyte ID No: 7478825CD1 <400> 17 Met Phe Ala Val His Leu Met Ala Phe Tyr Phe Ser Lys Leu Lys G1u Asp Gln Ile Lys Lys Val Asp Arg Phe Leu Tyr His Met Arg Leu Ser Asp Asp Thr Leu Leu Asp Ile Met Arg Arg Phe Arg Ala Glu Met Glu Lys Gly Leu Ala Lys Asp Thr,Asn Pro Thr Ala Ala Val Lys Met Leu Pro Thr Phe Val Arg Ala Ile Pro Asp Gly Ser Glu Asn Gly Glu Phe Leu Ser Leu Asp Leu Gly Gly Ser Lys Phe Arg Val Leu Lys Val Gln Val Ala Glu Glu Gly Lys Arg His Val Gln Met Glu Ser Gln Phe Tyr Pro Thr Pro Asn G1u Ile Ile Arg Gly Asn Gly Thr Glu Leu Phe Glu Tyr Val Ala Asp Cys Leu Ala Asp Phe Met Lys Thr Lys Asp Leu Lys His Lys Lys Leu Pro Leu Gly Leu Thr Phe Ser Phe Pro Cys Arg Gln Thr Lys Leu Glu Glu G1y Val Leu Leu Ser Trp Thr Lys Lys Phe Lys Ala Arg Gly Val Gln Asp Thr Asp Val Val Ser Arg Leu Thr Lys A1a Met Arg Arg His Lys Asp Met Asp Val Asp Ile Leu Ala Leu Val Asn Asp Thr Val Gly Thr Met Met Thr Cys Ala Tyr Asp Asp Pro Tyr Cys Glu Val Gly Val Ile Ile Gly Thr Gly Thr Asn Ala Cys Tyr Met Glu Asp Met Ser Asn Ile Asp Leu Val Glu Gly Asp Glu Gly Arg Met Cys Ile Asn Thr Glu Trp Gly Ala Phe Gly Asp Asp Gly Ala Leu Glu Asp Ile Arg Thr Glu Phe Asp Arg Glu Leu Asp Leu Gly Ser Leu Asn Pro Gly Lys Gln Leu Phe Glu Lys Met Ile Ser Gly Leu Tyr Leu Gly Glu Leu Val Arg Leu Ile Leu Leu Lys Met Ala Lys Ala Gly Leu Leu Phe Gly Gly Glu Lys Ser Ser Ala Leu His Thr Lys G1y Lys Ile Glu Thr Arg His Val Ala Ala.Met Glu Lys Tyr Lys Glu Gly Leu Ala Asn Thr Arg Glu I1e Leu Va1 Asp Leu Gly Leu Glu Pro Ser Glu Ala Asp Cys Ile Ala Val Gln His Val Cys Thr Ile Val Ser Phe Arg Ser Ala Asn Leu Cys Ala Ala Ala Leu Ala Ala Ile Leu Thr Arg Leu Arg Glu Asn Lys Lys VaI Glu Arg 395. 400 405 Leu Arg Thr Thr Val Gly Met Asp Gly Thr Leu Tyr Lys Ile His Pro Gln Tyr Pro Lys Arg Leu His Lys Val Val Arg Lys Leu Val Pro Ser Cys Asp Va1 Arg Phe Leu Leu Ser Glu Ser Gly Ser Thr Lys Gly Ala Ala Met Val Thr Ala Val Ala Ser Arg Val Gln Ala Gln Arg Lys Gln Ile Asp Arg Val Leu Ala Leu Phe Gln Leu Thr Arg Glu Gln Leu Val Asp Val Gln Ala Lys Met Arg Ala Glu Leu Glu Tyr Gly Leu Lys Lys Lys Ser His Gly Leu Ala Thr Val Arg Met Leu Pro Thr Tyr Val Cys Gly Leu Pro Asp Gly Thr Glu Lys Gly Lys Phe Leu Ala Leu Asp Leu Gly Gly Thr Asn Phe Arg Val Leu Leu Val Lys Ile Arg Ser Gly Arg Arg Ser Val Arg Met Tyr Asn Lys Ile Phe Ala Ile Pro Leu Glu Ile Met Gln Gly Thr Gly Glu Glu Leu Phe Asp His I1e Val Gln Cys Ile Ala Asp Phe Leu Asp Tyr Met Gly Leu Lys Gly Ala Ser Leu Pro Leu Gly Phe Thr Phe Ser Phe Pro Cys Arg Gln Met Ser Ile Asp Lys Gly Thr Leu Ile Gly Trp Thr Lys Gly Phe Lys Ala Thr Asp Cys Glu Gly Glu Asp Val Val Asp Met Leu Arg Glu Ala Ile Lys Arg Arg Asn G1u Phe Asp Leu Asp Ile Val Ala Val Val Asn Asp Thr Val Gly Thr Met Met Thr Cys Gly Tyr Glu Asp Pro Asn Cys Glu Ile Gly Leu Ile Ala Gly Thr Gly Ser Asn Met Cys Tyr Met Glu Asp Met Arg Asn Ile Glu Met Val Glu Gly Gly Glu G1y Lys Met Cys Ile Asn Thr Glu Trp Gly Gly Phe Gly Asp Asn Gly Cys Ile Asp Asp Ile Arg Thr Arg Tyr Asp Thr Glu Val Asp Glu Gly Ser Leu Asn Pro Gly Lys Gln Arg Tyr Glu Lys Met Thr Ser G1y Met Tyr Leu Gly Glu Ile Val Arg G1n Ile Leu Ile Asp Leu Thr Lys Gln Gly Leu Leu Phe Arg Gly Gln Ile Ser Glu Arg Leu Arg Thr Arg Gly Ile Phe Glu Thr Lys Phe Leu Ser Gln Ile Glu Ser Asp Arg Leu A1a Leu Leu Gln Val Arg Arg Ile Leu Gln Gln Leu Gly Leu Asp Ser Thr Cys Glu Asp Ser Ile Val Val Lys Glu Val Cys G1y A1a Val Ser Arg Arg Ala Ala Gln Leu Cys Gly A1a Gly Leu Ala Ala Ile Val Glu Lys Arg Arg Glu Asp Gln Gly Leu Glu His Leu Arg Ile Thr Val Gly Val Asp Gly Thr Leu Tyr Lys Leu His Pro His Phe Ser Arg Ile Leu Gln Glu Thr Val Lys G1u Leu A1a Pro Arg Cys Asp Val Thr Phe Met Leu Ser Glu Asp Gly Ser G1y Lys Gly Ala Ala Leu Ile Thr Ala Val Ala Lys Arg Leu Gln Gln Ala Gln Lys Glu Asn <210> 18 <211> 2380 <212> PRT
<213> Homo sapiens <220>
<221> misc feature <223> Incyte ID No: 7477141CD1 <400> 18 Met Asn His Pro Pro Trp Pro Ser Leu Asp Cys His Leu Lys Ala 1~ 5 10 15 Arg Ser Gly His Ala Leu Leu Ser Trp Pro Gly Gly Trp Ala Phe Pro Ile Ser Arg Glu Gln Asn Ala Ser Leu Ser Leu Cys Leu Ser Val Ser Leu Cys Val Arg Met Cys Val Ser Leu Thr Leu Cys Va1 Ser Ala Leu Cys Val Ala Pro Val Ala Ala Phe Pro Ser Ala His Pro Glu Ser Arg Ser Leu Ala Val Leu Ala Pro Leu Gln Asp Val Asp Val Gly Ala Gly Glu Met Ala Leu Phe Glu Cys Leu Val Ala Gly Pro Thr Asp Val Glu Val Asp Trp Leu Cys Arg Gly Arg Leu Leu Gln Pro A1a Leu Leu Lys Cys Lys Met His Phe Asp Gly Arg Lys Cys Lys Leu Leu Leu Thr Ser Val His Glu Asp Asp Ser Gly Val Tyr Thr Cys Lys Leu Ser Thr Ala Lys Asp Glu Leu Thr Cys Ser A1a Arg Leu Thr Val Arg Pro Ser Leu Ala Pro Leu Phe Thr Arg Leu Leu Glu Asp Val Glu Val Leu Glu Gly Arg Ala Ala Arg Phe Asp Cys Lys Ile Ser Gly Thr Pro Pro Pro Val Val Thr Trp Thr His Phe Gly Cys Pro Met Glu Glu Ser Glu Asn Leu Arg Leu Arg Gln Asp Gly Gly Leu His Ser Leu His Ile Ala His Val Gly Ser Glu Asp Glu Gly Leu Tyr Ala Val Ser Ala Val Asn Thr His Gly Gln Ala His Cys Ser Ala Gln Leu Tyr Val Glu Glu Pro Arg Thr Ala Ala Ser Gly Pro Ser Ser Lys Leu Glu Lys Met Pro Ser Ile Pro Glu Glu Pro Glu Gln Gly Glu Leu Glu Arg Leu Ser Ile Pro Asp Phe Leu Arg Pro Leu Gln Asp Leu Glu Va1 Gly Leu Ala Lys Glu Ala Met Leu Glu Cys Gln Val Thr Gly Leu Pro Tyr Pro Thr I1e Ser Trp Phe His Asn Gly His Arg Ile Gln Ser Ser Asp Asp Arg Arg Met Thr Gln Tyr Arg Asp Val His Arg Leu Val Phe Pro Ala Val Gly Pro Gln His Ala Gly Va1 Tyr Lys Ser Val Ile Ala Asn Lys Leu Gly Lys Ala Ala Cys Tyr Ala His Leu Tyr Val Thr Asp Val Val Pro Gly Pro Pro Asp Gly Ala Pro Gln Val Val Ala Val Thr Gly Arg Met Val Thr Leu Thr Trp Asn Pro Pro Arg Ser Leu Asp Met Ala Ile Asp Pro Asp Ser Leu Thr Tyr Thr Val Gln His Gln Val Leu Gly Ser Asp Gln Trp Thr Ala Leu Val Thr Gly Leu Arg Glu Pro Gly Trp Ala Ala Thr Gly Leu Arg Lys Gly Val Gln His Ile Phe Arg Val Leu Ser Thr Thr Val Lys Ser Ser Ser Lys Pro Ser Pro Pro Ser Glu Pro~Val Gln Leu Leu Glu His Gly Pro Thr Leu Glu Glu Ala Pro Ala Met Leu Asp Lys Pro Asp Ile Val Tyr Val Val Glu Gly Gln Pro Ala Ser Val Thr Val Thr Phe Asn His Val Glu Ala Gln Val Val Trp Arg Ser Cys Arg Gly Ala Leu Leu Glu Ala Arg Ala Gly Val Tyr Glu Leu Ser Gln Pro Asp Asp Asp Gln Tyr Cys Leu Arg I1e Cys Arg Val Ser Arg Arg Asp Met Gly Ala Leu Thr Cys Thr Ala Arg Asn Arg His Gly Thr G1n Thr Cys Ser Val Thr Leu Glu Leu Ala Glu Ala Pro Arg Phe Glu Ser Ile Met Glu Asp Val Glu Val Gly A1a Gly Glu Thr Ala Arg Phe Ala Val Val Val Glu Gly Lys Pro Leu Pro Asp Ile Met Trp Tyr Lys Asp Glu Val Leu Leu Thr Glu Ser Ser His Val Ser Phe Val Tyr Glu Glu Asn Glu Cys Ser Leu Val Val Leu Ser Thr Gly Ala Gln Asp Gly Gly Val Tyr Thr Cys Thr Ala Gln Asn Leu Ala Gly Glu Val Ser Cys Lys Ala Glu Leu Ala Va1 His Ser Ala Gln Thr Ala Met Glu Val Glu Gly Va1 Gly Glu Asp G1u Asp His Arg Gly Arg Arg Leu Ser Asp Phe Tyr Asp Ile His Gln Glu Ile Gly Arg Gly Ala Phe Ser Tyr Leu Arg Arg Ile Val Glu Arg Ser Ser G1y Leu Glu Phe Ala Ala Lys Phe Ile Pro Ser Gln Ala Lys Pro Lys Ala Ser Ala Arg Arg Glu Ala Arg Leu Leu Ala Arg Leu Gln His Asp Cys Val Leu Tyr Phe His Glu Ala Phe Glu Arg Arg Arg Gly Leu Val Ile Val Thr Glu Leu Cys Thr Glu Glu Leu Leu Glu Arg I1e Ala Arg Lys Pro Thr Val Cys Glu Ser Glu Ile Arg Ala Tyr Met Arg Gln Val Leu Glu Gly Ile His Tyr Leu His Gln Ser His Val Leu His Leu Asp Val Lys Pro Glu Asn Leu Leu Val Trp Asp Gly Ala Ala Gly Glu Gln Gln Val Arg Ile Cys Asp Phe Gly Asn Ala Gln Glu Leu Thr Pro Gly Glu Pro Gln Tyr Cys Gln Tyr Gly Thr Pro Glu Phe Val Ala Pro Glu Ile Val Asn Gln Ser Pro Val Ser Gly Val Thr Asp Ile Trp Pro Val Gly Val Val Ala Phe Leu Cys Leu Thr Gly Ile Ser Pro Phe Val G1y Glu Asn Asp Arg Thr Thr Leu Met Asn Ile Arg Asn Tyr Asn Val Ala Phe Glu Glu Thr Thr Phe Leu Ser Leu Ser Arg Glu Aha Arg Gly Phe Leu Ile Lys Val Leu Val Gln Asp Arg Leu Arg Pro Thr Ala Glu Glu Thr Leu Glu His Pro Trp Phe Lys Thr Gln Ala Lys Gly A1a Glu Val Ser Thr Asp His Leu Lys Leu Phe Leu Ser Arg Arg Arg Trp Gln Arg Ser Gln Ile Ser Tyr Lys Cys His Leu Val Leu Arg Pro Ile Pro Glu Leu Leu Arg Ala Pro Pro Glu Arg Val Trp Val Thr Met Pro Arg Arg Pro Pro Pro Ser Gly Gly Leu Ser Ser Ser Ser Asp Ser Glu Glu Glu Glu Leu Glu Glu Leu Pro Ser Val Pro Arg Pro Leu Gln Pro Glu Phe Ser G1y Ser Arg Val Ser Leu Thr Asp Ile Pro Thr Glu Asp Glu Ala Leu Gly Thr Pro Glu Thr Gly Ala Ala Thr Pro Met Asp Trp Gln Glu Gln Gly Arg Ala Pro Ser Gln Asp Gln Glu Ala Pro Ser Pro Glu Ala Leu Pro Ser Pro Gly Gln Glu Pro Ala Ala Gly Ala Ser Pro Arg Arg Gly Glu Leu Arg Arg Gly Ser Ser Ala Glu Ser Ala Leu Pro Arg A1a Gly Pro Arg Glu Leu Gly Arg Gly Leu His Lys Ala Ala Ser Val Glu Leu Pro Gln Arg Arg Ser Pro Gly Pro G1y Ala Thr Arg Leu Ala Arg Gly Gly Leu Gly Glu Gly Glu Tyr Ala Gln Arg Leu Gln Ala Leu Arg Gln Arg Leu Leu Arg Gly Gly Pro GIu Asp Gly Lys Val Ser Gly Leu Arg Gly Pro Leu Leu Glu Ser Leu Gly Gly Arg Ala Arg Asp Pro Arg Met Ala Arg Ala Ala Ser Ser Glu A1a Ala Pro His His Gln Pro Pro Leu Glu Asn Arg Gly Leu Gln Lys Ser Ser Ser Phe Ser Gln Gly Glu A1a Glu Pro Arg G1y Arg His Arg Arg Ala G1y Ala Pro Leu Glu Ile Pro Val Ala Arg Leu Gly Ala Arg Arg Leu Gln Glu Ser Pro Ser Leu Ser Ala Leu Ser Glu Ala Gln Pro Ser Ser Pro Ala Arg Pro Ser Ala Pro Lys Pro Ser Thr Pro Lys Ser Ala Glu Pro Ser Ala Thr Thr Pro Ser Asp Ala Pro Gln Pro Pro Ala Pro G1n Pro Ala Gln Asp Lys Ala~Pro Glu Pro Arg Pro Glu Pro Val Arg Ala Ser Lys Pro Ala Pro Pro Pro Gln Ala Leu Gln Thr Leu Ala Leu Pro Leu Thr Pro Tyr Ala Gln Ile Ile Gln Ser Leu Gln Leu Ser Gly His Ala Gln Gly Pro Ser Gln Gly Pro Ala Ala Pro Pro Ser Glu Pro Lys Pro His Ala Ala Val Phe Ala Arg Val Ala Ser Pro Pro Pro Gly Ala Pro Glu Lys Arg Val Pro Ser Ala Gly Gly Pro Pro Val Leu Ala Glu Lys Ala Arg Val Pro Thr Val Pro Pro Arg Pro Gly Ser Ser Leu Ser Ser Ser Ile Glu Asn Leu 1430 1435' 1440 Glu Ser Glu Ala Val Phe Glu Ala Lys Phe Lys Arg Ser Arg Glu Ser Pro Leu Ser Leu Gly Leu Arg Leu Leu Ser Arg Ser Arg Ser Glu Glu Arg Gly Pro Phe Arg Gly Ala Glu G1u Glu Asp G1y Ile Tyr Arg Pro Ser Pro Ala Gly Thr Pro Leu Glu Leu Val Arg Arg Pro Glu Arg Ser Arg Ser Val Gln Asp Leu Arg Ala Val Gly Glu Pro Gly Leu Val Arg Arg Leu Ser Leu Ser Leu Ser Gln Arg Leu Arg Arg Thr Pro Pro Ala Gln Arg His Pro Ala Trp Glu Ala Arg Gly Gly Asp Gly Glu Ser Ser Glu Gly Gly Ser Ser Ala Arg Gly Ser Pro Val Leu Ala Met Arg Arg Arg Leu Ser Phe Thr Leu Glu Arg Leu Ser Ser Arg Leu Gln Arg Ser Gly Ser Ser Glu Asp Ser 1580 1585 ' 1590 Gly Gly Ala Ser Gly Arg Ser Thr Pro Leu Phe Gly Arg Leu Arg Arg Ala Thr Ser Glu Gly Glu Ser Leu Arg Arg Leu Gly Leu Pro His Asn Gln Leu Ala Ala Gln Ala Gly Ala Thr Thr Pro Ser Ala Glu Ser Leu Gly Ser Glu Ala Ser Ala Thr Ser Gly Ser Ser Ala Pro Gly Glu Ser Arg Ser Arg Leu Arg Trp G1y Phe Ser Arg Pro Arg Lys Asp Lys Gly Leu Ser Pro Pro Asn Leu Ser Ala Ser Val Gln Glu Glu Leu Gly His Gln Tyr Val Arg Ser Glu Ser Asp Phe Pro Pro Val Phe His Ile Lys Leu Lys Asp Gln Va1 Leu Leu Glu Gly Glu A1a Ala Thr Leu Leu Cys Leu Pro Ala Ala Cys Pro Ala Pro His Ile Ser Trp Met Lys Asp Lys Lys Ser Leu Arg Ser G1u Pro Ser Val Ile Ile Val Ser Cys Lys Asp Gly Arg Gln Leu Leu Ser Ile Pro Arg Ala Gly Lys Arg His A1a Gly Leu Tyr Glu Cys Ser Ala Thr Asn Val Leu Gly Ser Ile Thr Ser Ser Cys Thr Val Ala Val Ala Arg Val Pro Gly Lys Leu Ala Pro Pro Glu Val Pro Gln Thr Tyr Gln Asp Thr Ala Leu Val Leu Trp Lys Pro Gly Asp Ser Arg Ala Pro Cys Thr Tyr Thr Leu Glu Arg Arg Val Asp Gly Glu Ser Val Trp His Pro Val Ser Ser Gly Ile Pro Asp Cys Tyr Tyr Asn Va1 Thr His Leu Pro Val Gly Va1 Thr Val Arg Phe Arg Val Ala Cys Ala Asn Arg Ala Gly Gln Gly Pro Phe Ser Asn Ser Ser Glu Lys Val Phe Val Arg Gly Thr Gln Asp Ser Ser Ala Val Pro Ser Ala Ala His Gln Glu Ala Pro Val Thr Ser Arg Pro Ala Arg Ala Arg Pro Pro Asp Ser Pro Thr Ser Leu Ala Pro Pro Leu Ala Pro Ala Ala Pro Thr Pro Pro Ser Val Thr Val Ser Pro Ser Ser Pro Pro Thr Pro Pro Ser Gln Ala Leu Ser Ser Leu Lys Ala Val Gly Pro Pro Pro Gln Thr Pro Pro Arg Arg His Arg Gly Leu Gln Ala Ala Arg Pro Ala Glu Pro Thr Leu Pro Ser Thr His Val Thr Pro Ser Glu Pro Lys Pro Phe Val Leu Asp Thr Gly Thr Pro Ile Pro A1a Ser Thr Pro Gln Gly Val Lys Pro Val Ser Ser Ser Thr Pro Val Tyr Val Val Thr Ser Phe Val Ser Ala Pro Pro Ala Pro Glu Pro Pro Ala Pro Glu Pro Pro Pro Glu Pro Thr Lys Val Thr Val Gln Ser Leu Ser Pro Ala Lys Glu Val Val Ser Ser Pro Gly Ser Ser Pro Arg Ser Ser Pro Arg Pro Glu Gly Thr Thr Leu Arg Gln Gly Pro Pro Gln Lys Pro Tyr Thr Phe Leu Glu Glu Lys Ala Arg Gly Arg Phe Gly Val Val Arg Ala Cys Arg Glu Asn Ala Thr Gly Arg Thr Phe Val Ala Lys Ile Val Pro Tyr Ala Ala Glu Gly Lys Arg Arg Val Leu Gln Glu Tyr Glu Val Leu Arg Thr Leu His His Glu Arg Ile Met Ser Leu His Glu Ala Tyr Ile Thr Pro 2135 2140 ~ 2145 Arg Tyr Leu Val Leu Ile Ala Glu Ser Cys Gly Asn Arg Glu Leu Leu Cys Gly Leu Ser Asp Arg Phe Arg Tyr Ser Glu Asp Asp Val Ala Thr Tyr Met Val Gln Leu Leu Gln Gly Leu Asp Tyr Leu His Gly'His His Val Leu His Leu Asp Ile Lys Pro Asp Asn Leu Leu Leu Ala Pro Asp Asn Ala Leu Lys Ile Val Asp Phe Gly Ser Ala Gln Pro Tyr Asn Pro Gln Ala Leu Arg Pro Leu Gly His Arg Thr Gly Thr Leu Glu Phe Met Ala Pro Glu Met Val Lys Gly Glu Pro Ile Gly Ser Ala Thr Asp Ile Trp Gly Ala Gly Val Leu Thr Tyr Ile Met Leu Ser Gly Arg Ser Pro Phe Tyr G1u Pro Asp Pro Gln Glu Thr Glu Ala Arg Ile Val Gly Gly Arg Phe Asp Ala Phe Gln Leu Tyr Pro Asn Thr Ser Gln Ser Ala Thr Leu Phe Leu Arg Lys Val Leu Ser Val His Pro Trp Ser Arg Pro Ser Leu Gln Asp Cys Leu Ala His Pro Trp Leu Gln Asp Ala Tyr Leu Met Lys Leu Arg Arg Gln Thr Leu Thr Phe Thr Thr Asn Arg Leu Lys Glu Phe Leu Gly Glu Gln Arg Arg Arg Arg Ala Glu Ala Ala Thr Arg His Lys Val Leu Leu Arg Ser Tyr Pro Gly Gly Pro <210> 19 <211> 505 <212> PRT
<213> Homo Sapiens <220>
<221> misc feature <223> Incyte ID No: 2190612CD1 <400> 19 Met Glu Gly Gly Pro Ala Val Cys Cys Gln Asp Pro Arg Ala Glu Leu Val Glu Arg Val Ala Ala Ile Asp Val Thr His Leu Glu Glu Ala Asp Gly Gly Pro Glu Pro Thr Arg Asn Gly Val Asp Pro Pro Pro Arg Ala Arg Ala Ala Ser Val Ile Pro Gly Ser Thr Ser Arg Leu Leu Pro Ala Arg Pro Ser Leu Ser Ala Arg Lys Leu Ser Leu Gln Glu Arg Pro Ala Gly Ser Tyr Leu Glu Ala Gln AIa Gly Pro Tyr Ala Thr Gly Pro Ala Ser His I1e Ser Pro Arg Ala Trp Arg Arg Pro Thr I1e Glu Ser His His Val Ala Ile Ser Asp Ala Glu 110 1~.5 220 Asp Cys Val Gln Leu Asn Gln Tyr Lys Leu Gln Ser Glu Ile Gly Lys G1y Ala Tyr G1y Val Va1 Arg Leu Ala Tyr Asn Glu Ser Glu Asp Arg His Tyr Ala Met Lys Val Leu Ser Lys Lys Lys Leu Leu Lys Gln Tyr Gly Phe Pro Arg Arg Pro Pro Pro Arg Gly Ser Gln Ala Ala Gln Gly Gly Pro Ala Lys Gln Leu Leu Pro Leu Glu Arg Val Tyr Gln Glu Ile Ala Ile Leu Lys Lys Leu Asp His Val Asn 200 205. 210 Val Va1 Lys Leu Ile Glu Val Leu Asp Asp Pro Ala Glu Asp Asn Leu Tyr Leu Val Phe Asp Leu Leu Arg Lys Gly Pro Va1 Met Glu Val Pro Cys Asp Lys Pro Phe Ser Glu Glu Gln Ala Arg Leu Tyr Leu Arg Asp Val I1e Leu G1y Leu Glu Tyr Leu His Cys Gln Lys Ile Val His Arg Asp Ile Lys Pro Ser Asn Leu Leu Leu Gly Asp Asp Gly His Val Lys Ile Ala Asp Phe Gly Val Ser Asn Gln Phe Glu Gly Asn Asp Ala Gln Leu Ser Ser Thr Ala Gly Thr Pro Ala Phe Met Ala Pro Glu Ala Ile Ser Asp Ser Gly Gln Ser Phe Ser G1y Lys Ala Leu Asp Val Trp Ala Thr Gly Val Thr Leu Tyr Cys Phe Val Tyr Gly Lys Cys Pro Phe Ile Asp Asp Phe Ile Leu Ala Leu His Arg Lys Ile Lys Asn Glu Pro Val Val Phe Pro G1u Glu Pro Glu Ile Ser Glu Glu Leu Lys Asp Leu Ile Leu Lys Met Leu Asp Lys Asn Pro Glu Thr Arg Ile Gly Va1 Pro Asp Ile Lys Leu His Pro Trp Val Thr Lys Asn Gly Glu Glu Pro Leu Pro Ser G1u Glu Glu His Cys Ser Val Val Glu Val Thr Glu Glu Glu Val Lys Asn Ser Val Arg Leu Ile Pro Ser Trp Thr Thr Val I1e Leu Val Lys Ser Met Leu Arg Lys Arg Ser Phe Gly Asn Pro Phe Glu Pro Gln Ala Arg Arg Glu Glu Arg Ser Met Ser Ala Pro Gly Asn Leu Leu Val Lys Glu Gly Phe Gly Glu Gly Gly Lys Ser Pro Glu Leu Pro G1y Val Gln Glu Asp Glu Ala Ala Ser <210> 20 <211> 1572 <212> PRT
<213> Homo sapiens <220>
<221> misc feature <223> Incyte ID No: 7477549CD1 <400> 20 Met Glu Arg Arg Leu Arg Ala Leu Glu Gln Leu Ala Arg Gly Glu Ala Gly Gly Cys Pro Gly Leu Asp Gly Leu Leu Asp Leu Leu Leu Ala Leu His His Glu Leu Ser Ser G1y Pro Leu Arg Arg Glu Arg Ser Val Ala G1n Phe Leu Ser Trp Ala Ser Pro Phe Val Ser Lys Val Lys Glu Leu Arg Leu G1n Arg Asp Asp Phe Glu Ile Leu Lys Va1 Ile Gly Arg Gly A1a Phe Gly G1u Val Thr Val Val Arg Gln Arg Asp Thr Gly Gln Ile Phe Ala Met Lys Met Leu His Lys Trp Glu Met Leu Lys Arg Ala Glu Thr Ala Cys Phe Arg Glu Glu Arg Asp Val Leu Val Lys Gly Asp Ser Arg Trp Val Thr Thr Leu His Tyr Ala Phe Gln Asp Glu Glu Tyr Leu Tyr Leu Val Met Asp Tyr Tyr Ala Gly Gly Asp Leu Leu Thr Leu Leu Ser Arg Phe Glu Asp Arg Leu Pro Pro Glu Leu Ala Gln Phe Tyr Leu Ala Glu Met Val Leu Ala Ile His Ser Leu His Gln Leu Gly Tyr Val His Arg Asp Val Lys Pro Asp Asn Val Leu Leu Asp Val Asn Gly His Ile Arg Leu Ala Asp Phe Gly Ser Cys Leu Arg Leu Asn Thr Asn Gly Met Val Asp Ser Ser Val Ala Val Gly Thr Pro Asp Tyr Ile Ser Pro Glu Ile Leu Gln Ala Met Glu Glu Gly Lys Gly His Tyr Gly Pro Gln Cys Asp Trp Trp Ser Leu Gly Val Cys Ala Tyr Glu Leu Leu Phe Gly Glu Thr Pro Phe Tyr Ala Glu Ser Leu Val Glu Thr Tyr Gly Lys Ile Met Asn His Glu Asp His Leu Gln Phe Pro Pro Asp Val Pro Asp Val Pro Ala Ser Ala Glri Asp Leu Ile Arg Gln Leu Leu Cys Arg Gln Glu Glu Arg Leu Gly Arg Gly Gly Leu Asp Asp Phe Arg Asn His Pro Phe Phe Glu Gly Val Asp Trp Glu Arg Leu Ala Ser Ser Thr Ala Pro Tyr Ile Pro Glu Leu Arg Gly Pro Met Asp Thr Ser Asn Phe Asp Val Asp Asp Asp Thr Leu Asn His Pro Gly Thr Leu Pro Pro Pro Ser His Gly Ala Phe Ser Gly His His Leu Pro Phe Val Gly Phe Thr Tyr Thr Ser Gly Ser His Ser Pro Glu Ser Ser Sex G1u Ala Trp Ala Ala Leu Glu Arg Lys Leu Gln Cys Leu Glu Gln Glu Lys Val Glu Leu Ser Arg Lys His Gln Glu Ala Leu His Ala Pro Thr Asp His Arg Glu Leu Glu Gln Leu Arg Lys Glu Val Gln Thr Leu Arg Asp Arg Leu Pro Glu Met Leu Arg Asp Lys Ala Ser Leu Ser Gln Thr Asp Gly Pro Pro Ala Gly Ser Pro Gly Gln Asp Ser Asp Leu Arg Glri Glu Leu Asp Arg Leu His Arg Glu Leu Ala Glu Gly Arg Ala Gly Leu Gln Ala Gln Glu Gln Glu Leu Cys Arg Ala Gln Gly Gln Glri G1u Glu Leu Leu Gln Arg Leu Gln Glu Ala Gln Glu Arg Glu Ala Ala Thr Ala Ser Gln Thr Arg Ala Leu Ser Ser Gln Leu Glu Glu Ala Arg Ala Ala Gln Arg Glu Leu Glu Ala Gln Val Ser Ser Leu Ser Arg Gln Val Thr Gln Leu Gln Gly Gln Trp Glu G1n Arg Leu Glu Glu Ser Ser Glri Ala Lys Thr Ile His Thr Ala Ser Glu Thr Asn Gly Met Gly Pro Pro 590 , 595 , 600 Glu Gly Gly Pro Gln Glu Ala Gln Leu Arg Lys G1u Val Ala Ala Leu Arg Glu Gln Leu Glu Gln Ala His Ser His Arg Pro Ser Gly Lys Glu Glu Ala Leu Cys Gln Leu G1n Glu Glu Asn Arg Arg Leu Ser Arg Glu Gln Glu Arg Leu Glu Ala G1u Leu Ala Gln Glu Gln Glu Ser Lys Gln Arg Leu Glu Gly G1u Arg Arg Glu Thr Glu Ser Asn Trp Glu Ala Gln Leu Ala Asp Ile Leu Ser Trp Va1 Asn Asp Glu Lys Val Ser Arg Gly Tyr Leu Gln Ala Leu Ala Thr Lys Met Ala Glu Glu Leu G1u Ser Leu Arg Asn Val Gly Thr Gln Thr Leu Pro Ala Arg Pro Leu Lys Met Glu Ala Ser Ala Arg Leu Glu Leu Gln Ser Ala Leu Glu Ala Glu Ile Arg Ala Lys Gln Gly Leu Gln Glu Arg Leu Thr Gln Val Gln Glu Ala Gln Leu Gln Ala Glu Arg Arg Leu Gln Glu Ala Glu Lys Gln Ser Gln Ala Leu Gln Gln Glu Leu Ala Met Leu Arg Glu G1u Leu Arg Ala Arg Gly Pro Val Asp Thr Lys Pro Ser Asn Ser Leu Ile Pro Phe Leu Ser Phe Arg Ser Ser Glu Lys Asp Ser Ala Lys Asp Pro G1y I1e Ser Gly Glu Ala Thr Arg His Gly Gly Glu Pro Asp Leu Arg Pro Glu Gly Arg Arg Ser Leu Arg Met Gly Ala Val Phe Pro Arg Ala Pro Thr Ala Asn Thr Ala Ser Thr Glu Gly Leu Pro A1a Lys Gly Trp Gly Met Gly Pro Trp Glu Ala Leu Gly Asn Gly Cys Pro Pro Pro Gln Pro Gly Ser His Thr Leu Arg Pro Arg Ser Phe Pro Sex Pro Thr Lys Cys Leu Arg Cys Thr Ser Leu Met Leu Gly Leu Gly Arg Gln G1y Leu Gly Cys Asp Ala Cys Gly Tyr Phe Cys His Thr Thr Cys Ala Pro Gln Ala Pro Pro Cys Pro Val Pro Pro Asp Leu Leu Arg Thr Ala Leu Gly Val His Pro Glu Thr Gly Thr Gly Thr Ala Tyr Glu Gly Phe Leu Ser Val Pro Arg Pro Ser G1y Val Arg Arg Gly Trp Gln Arg Val Phe Ala Ala Leu Ser Asp Ser Arg Leu Leu Leu Phe Asp Ala Pro Asp Leu Arg Leu Ser Pro Pro Ser Gly Ala Leu Leu Gln Val Leu Asp Leu Arg Asp Pro Gln Phe Ser Ala Thr Pro Val Leu A1a Ser Asp Val Ile His Ala Gln Ser Arg Asp Leu Pro Arg Ile Phe Arg Val Thr Thr Ser Gln Leu Ala Val Pro Pro Thr Thr Cys Thr Val Leu Leu Leu Ala Glu Ser Glu Gly Glu Arg Glu Arg Trp Leu Gln Val Leu Gly Glu Leu Gln Arg Leu Leu Leu Asp Ala Arg Pro Arg Pro Arg Pro Va1 Tyr Thr Leu Lys Glu Ala Tyr Asp Asn Gly Leu Pro Leu Leu Pro His Thr Leu Cys Ala Ala Ile Leu Asp Gln Asp Arg Leu Ala Leu Gly Thr Glu Glu Gly Leu Phe Val Ile His Leu Arg Ser Asn Asp Ile Phe Gln Val Gly Glu Cys Arg Arg Val Gln Gln Leu Thr Leu Ser Pro Ser Ala Gly Leu Leu Val Va1 Leu Cys Gly Arg Gly Pro Ser Val Arg Leu Phe Ala Leu Ala Glu Leu Glu Asn Ile Glu Val Ala Gly Ala Lys Ile Pro Glu Ser Arg Gly Cys Gln Val Leu Ala Ala Gly Ser Ile Leu Gln Ala Arg Thr Pro Val Leu Cys Val Ala Val Lys Arg Gln Val Leu Cys Tyr Gln Leu Gly Pro Gly Pro Gly Pro Trp Gln Arg Arg Ile Arg Glu Leu Gln Ala Pro Ala Thr Val Gln Ser Leu Gly Leu Leu Gly Asp Arg Leu Cys Val Gly Ala Ala G1y G1y Phe Ala Leu Tyr Pro Leu Leu Asn Glu Ala Ala Pro Leu Ala Leu Gly Ala Gly Leu Val Pro Glu Glu Leu Pro Pro Ser Arg Gly Gly Leu Gly Glu Ala Leu Gly Ala Val Glu Leu Ser Leu Ser Glu Phe Leu Leu Leu Phe Thr Thr Ala Gly Ile Tyr Val Asp Gly Ala Gly Arg Lys Ser Arg Gly His Glu Leu Leu Trp Pro Ala Ala Pro Met Gly Trp Gly Tyr A1a AIa Pro Tyr Leu Thr Val Phe Ser Glu Asn Ser Ile Asp Val Phe Asp Val Arg Arg Ala Glu Trp Val Gln Thr Val Pro Leu Lys Lys Val Arg Pro Leu Asn Pro Glu Gly Ser Leu Phe Leu Tyr Gly Thr Glu Lys Val Arg Leu Thr Tyr Leu Arg Asn Gln Leu Ala Glu Lys Asp Glu Phe Asp Ile Pro Asp Leu Thr Asp Asn Ser Arg Arg Gln Leu Phe Arg Thr Lys Ser Lys Arg Arg Phe Phe Phe Arg Val Ser Glu Glu Gln Gln Lys Gln Gln Arg Arg Glu Met Leu Lys Asp Pro Phe Val Arg Ser Lys Leu Ile Ser Pro Pro Thr Asn Phe Asn His Leu Val His Va1 Gly Pro Ala Asn G1y Arg Pro Gly Ala Arg Asp Lys Ser 1460 1465 ' 1470 Pro Ser Gln Pro Leu Arg Thr Val Thr Gln Gln Ala Pro Glu Glu 1475 1480 . 1485 Lys Gly Arg Val Ala Arg Gly Ser Gly Pro Gln Arg Pro His Ser Phe Ser Glu Ala Leu Arg Arg Pro Ala Ser Met Gly Ser Glu Gly Leu Gly Gly Asp Ala Asp Pro Thr Gly Ala Val Lys Arg Lys Pro Trp Thr Ser Leu Ser Ser Glu Ser Val Ser Cys Pro Gln Gly Ser Leu Ser Pro Ala Thr Ser Leu Met G1n Val Ser Glu Arg Pro Arg Ser Leu Pro Leu Ser Pro Glu Leu Glu Ser Ser Pro <210> 21 <211> 4298 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte TD No: 2564295CB1 <400> 21 gccactgaga ggcccaccag gtcccttctt ctcggatctg gcagaccaag gactagcgta 60 cggacctgcg cttaggggtc tccaagagga caaggagcct cctaggagct gagaggggct 120 cccagaggcc aaggctgtcc~acgttctccc gggtcgaggc tgccagaagt taccccgcag 180 atgtcgaggc accgggaagc agaatctaga cacagcgctc cccagaagcc cgggcgcgct 240 ggctgcccct ccggcggtgc agccccactt ggaagaagcc tgtgggctta tcacaccgtt 300 ctccccagag tcaccgggag gagagccggg actggacaca agccagggct gggacaatgg 360 cagtgcctag tctgtggccc tggggagcat gcctgcctgt gatcttcctc tccttgggat 420 ttggcctgga tacagtagag gtgtgcccca gcctggatat tcgctcagag gtggcagagc 480 ttcgtcagct ggagaactgc agcgtggtgg agggccacct gcagatcctg ctcatgttca 540 cagccaccgg,ggaggacttc cgcggcctca gcttccctcg cctcacccag gtcaccgact 600 acctgctgct cttccgtgtc tacggactgg agagcctgcg cgacctcttc cccaacctag 660 cagtcatccg cgggacgcgc ctcttcctgg gctatgcact ggtcatcttt gagatgccac 720 atctgcgtga ~cgtggcactg cctgcacttg gggccgtgct gcgtggggct gtgcgtgtgg 780 agaagaacca ggagctctgc cacctctcca ccattgactg gggactgctg cagccagcac 840 ctggcgccaa ccacatcgtg ggcaacaagc tgggcgagga gtgtgctgac gtgtgccctg 900 gtgtgctggg tgctgctggt gagccctgtg ccaagaccac cttcagcggg cacactgact 960 acagatgctg gacctccagc cactgccaga gagtgtgccc ctgcccccat gggatggctt 1020 gcacagcgag gggcgagtgc tgccacaccg aatgcctggg gggctgcagc cagccagaag 1080 accctcgtgc ctgtgtagct tgccgccacc tctacttcca gggtgcctgc ctgtgggcct 1140 gcccgccagg cacctaccag tatgagtcct ggcgctgtgt cacagctgag cgctgtgcca 1200 gcctgcactc tgtgcccggc cgtgcctcca ccttcggcat acaccagggc agttgcctgg 1260 cccagtgccc ttctggcttc acccgtaata gcagcagcat attctgccac aagtgcgagg 1320 ggctgtgccc taaagagtgc aaggtaggca ccaagaccat cgactccatc caggcggcac 1380 aggatcttgt gggctgcacg catgtggagg gaagcctcat cctcaacctt cgccagggct 1440 acaacctgga gccacagctg cagcacagcc tggggctggt agaaaccatt actggcttcc 1500 tcaaaatcaa gcactccttt gccctcgtgt ccctgggctt tttcaagaac ctcaaactaa 1560 tccggggaga cgccatggtg gatgggaact acactctcta cgtgctggac aaccagaacc 1620 tacaacagct agggtcctgg gtggccgcgg ggctcaccat tcccgtgggc aagatctact 1680 tcgccttcaa cccgcgcctc tgcttggaac acatctaccg actggaggag gtgacaggca 1740 cgcgaggtcg gcagaacaag gctgagatca acccccgcac caacggagac cgcgccgcct 1800 gccagactcg caccctgcgc ttcgtgtcca acgtgacgga ggcagaccgc atcctgctac 1860 gctgggagcg ctatgagcca ctggaggccc gcgacctgct cagcttcatc gtgtactaca 1920 aggagtcccc attccagaac gccacagagc acgtgggtcc agatgcttgt ggaacccaga 1980 gctggaacct gctggatgtg gagctgcccc taagccgcac ccaggagcca ggggtgaccc 2040 tagcctccct caagccttgg acacagtacg cagtgtttgt gcgggccatc acgctaacca 2100 ctgaggagga cagccctcat caaggagccc agagtcccat cgtctacctc cgaacgctgc 2160 ctgcagctcc cacggtgccc caagacgtca tctccacgtc caactcctcc tcccacctcc 2220 tggtgcgctg gaagccaccg acccagcgca atgggaacct cacctactac ctggtgctgt 2280 ggcagcggct ggcagaggac ggcgacctct acctcaatga ctactgccac cgcggcttgc 2340 ggctgcccac cagcaacaac gatccgcgct tcgacggcga agacggggat cctgaggccg 2400 agatggagtc cgactgctgc ccttgccagc acccacctcc tggtcaggtt ctgcccccgc 2460 tggaggcgca agaggcctcg ttccagaaga agtttgaaaa ctttctacac aacgcgatca 2520 ccatccccat atccccttgg aaggtgacgt ccatcaacaa gagcccccaa agggactcag 2580 ggcggcaccg ccgggcagct gggcccctcc ggctgggggg caacagctcg gatttcgaga 2640 tccaggagga caaggtgccc cgtgagcgag cggtgctgag cggcctgcgc cacttcacgg 2700 aataccggat cgacatccat gcctgcaacc acgcggcgca caccgtgggc tgcagcgccg 2760 ccaccttcgt ctttgcgcgc accatgcccc acagagaggc tgatggtatt ccaggaaagg 2820 tggcctggga ggcctccagc aagaacagtg tccttctgcg ctggctcgag ccaccagacc 2880 ccaacggact catcctcaag tacgaaatca agtaccgccg cttgggagag gaggccacag 2940 tgctgtgtgt gtcccgtctt cgatatgcga agtttggggg agtccacctg gccctgctgc 3000 cccctggaaa ctactctgcc agggttaggg caacctcact ggctggcaat ggctcttgga 3060 cagacagtgt tgccttctac atccttggcc cagaggagga ggatgctggg gggctgcatg 3120 tcctcctcac tgccacccct gtggggctca cgctgctcat cgttcttgct gcccttggtt 3180 tcttctacgg caagaagaga aacagaaccc tgtatgcttc tgtgaatcca gagtacttca 3240 gcgcctctga tatgtatgtc cctgatgaat gggaggtgcc tcgggagcag atctcgataa 3300 tccgggaact gggccagggc tcttttggga tggtatatga ggggctggca cgaggacttg 3360 aggctggaga ggagtccaca cccgtggccc tgaagacggt gaatgagctg gccagcccac 3420 gggaatgcat tgagttcctc aaggaagctt ctgtcatgaa agccttcaag tgtcaccatg 3480 tggtgcgtct cctgggtgtg gtatctcagg gccagccaac tctggtcatc atggagttaa 3540 tgacccgtgg ggacctcaag agccatcttc gatctttgcg gcctgaggca gagaacaacc 3600 ctgggctccc acagccagca ttgggggaaa tgatccaaat ggctggtgag attgcagacg 3660 gcatggccta ccttgctgcc aacaagtttg tgcaccgaga tctagcagcc cgcaactgca 3720 tggtgtccca ggacttcacc gtcaagatcg gggacttcgg gatgactcgg gacgtgtatg 3780 agacagacta ttaccgcaag ggtgggaagg ggctgctgcc cgtgcgctgg atggcccccg 3840 agtccctcaa agatgggatc ttcaccaccc actcggatgt ctggtccttt ggcgtggtac 3900 tctgggagat tgtgaccctg gcagaacaac cctaccaggg cctgtccaat gagcaggtgc 3960 tgaagttcgt catggatggc ggggtcctgg aggagctgga gggctgtccc cttcagctgc 4020 aggagctgat gagccgctgc tggcagccga acccacgcct gcgcccatct ttcacacaca 4080 ttctggacag catacaggag gagctgcggc cctccttccg cctcctctcc ttctactaca 4140 gcccggaatg ccggggggcc cggggctccc tgcctaccac cgatgcagag cctgactcct 4200 cacccactcc aagagactgc agccctcaaa atgggggtcc agggcactga ggggcacctc 4260 attccctggc tggcctccca tggggagaca ggaaggga 4298 <210> 22 <211> 2863 <212> DNA , <213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 2837050CB1 <400> 22 atgatggaag aattgcatag cctggaccca cgacggcagg aattattgga ggccaggttt 60 actagagtag gtgttagtaa gggaccactt aatagtgagt cttccaacca gagcttgtgc 120 agcgtcggat ccttgagtga taaagaagta gagactcccg agaaaaagca gaatgaccag 180 cgaaatcgga aaagaaaagc tgaaccatat gaaactagcc aagggaaagg cactcctagg 240 ggacataaaa ttagtgatta ctttgagcga cgagtagaac agcccctcta tggtttagat 300 ggcagtgctg caaaggaggc aacggaggag cagtctgctc tgccaaccct catgtcagtg 360 atgctagcaa aacctcggct tgacacagag cacgtggcgc aaaggggagc tggcctctgc 420 ttcacttttg tttcagctca gcaaaacagt ccctcatcta cgggatctgg caacacagag 480 cattcctgca gctcccaaaa acagatctcc atccagcaca gacagaccca gtccgacctc 540 acaatagaaa aaatatctgc actagaaaac agtaagaatt ctgacttaga gaagaaggag 600 ggaagaatag atgatttatt aagagccaac tgtgatttga gacggcagat tgatgaacag 660 caaaagatgc tagagaaata caaggaacga ttaaatagat gtgtgacaat gagcaagaaa 720 ctccttatag aaaagtcaaa acaagagaag atggcgtgta gagataagag catgcaagac 780 cgcttgagac tgggccactt tactactgtc cgacacggag cctcatttac tgaacagtgg 840 acagatggtt atgcttttca gaatcttatc aagcaacagg aaaggataaa ttcacagagg 900 gaagagatag aaagacaacg gaaaatgtta gcaaagcgga aacctcctgc catgggtcag 960 gcccctcctg caaccaatga gcagaaacag cggaaaagca agaccaatgg agctgaaaat 1020 gaaacgttaa cgttagcaga ataccatgaa caagaagaaa tcttcaaact cagattaggt 1080 catcttaaaa aggaggaagc agagatccag gcagagctgg agagactaga aagggttaga 1140 aatctacata tcagggaact aaaaaggata cataatgaag ataattcaca atttaaagat 1200 catccaacgc taaatgacag atatttgttg ttacatcttt tgggtagagg aggtttcagt 1260 gaagtttaca aggcatttga tctaacagag caaagatacg tagctgtgaa aattcaccag 1320 ttaaataaaa actggagaga tgagaaaaag gagaattacc acaagcatgc atgtagggaa 1380 taccggattc ataaagagct ggatcatccc agaatagtta agctgtatga ttacttttca 1440 ctggatactg actcgttttg tacagtatta gaatactgtg agggaaatga tctggacttc 1500 tacctgaaac agcacaaatt aatgtcagag aaagaggcct ggtccattat catgcagatt 1560 gtgaatgctt taaagtactt aaatgaaata aaacctccca tcatacacta tgacctcaaa 1620 ccaggtaata ttcttttagt aaatggtaca gtgtgtggag agagaaaaat tacagatttt 1680 ggtctttcga agatcatgga tgatgatagc tacaattcag tgggtggcat ggagctgaca 1740 tcacaaggtg ctggcactta ttggtattta ccaccggagt gttttgtggt tgagaaagaa 1800 ccaccaaaga tctcaaataa agttgatgtg tggtcggtgg gtgtgatctt ctatcagtgt 1860 ctttctggag ggaagccttt tggccataac cagtctcagc aagacatcct acaagagaat 1920 actattctta aagctgctga agtgcagttc ccgccaaagc cagtagtaac acctgaagca 1980 aaggcgttta ttcgacgatg cttggcctac cgaaaggagg actgcattga tgcccagcag 2040 ctggcctgtg atccctactt gttgcctcac atccgaaagt cagtctctac aagtagccct 2100 gctggagctg ctattgcatc aacctctggg gcgtccaata acagttcttc taattgagac 2160 tgactccaag gccacaaact gttcaacaca cacaaagtgg acaaatggcg ttcagcagcg 2220 ggtttggaac atagcgaatc tgaatggatc tgatgaaacc tgaaccaggt gcttttattt 2280 tcttgctttt ttcccatcca ctgagcatga cagcatggat tctctttaag gagaaacctt 2340 gggcagctcc agccaggcct cataggaaaa ggcccggcat gaggttctgg cgtcaatggc 2400 cactgtgtat ggctgctctg agtgaggaaa aaactaaaaa gaaaaactgg ttccatgtac 2460 tgtgaacttg aaaacatgca gactcacggg ggttcctgat gcaatgcttc agatgaagat 2520 tgtggacttg aaaatacaga ctagaaggcc gggcacagtg gctcatgcct gtaatctcag 2580 cactttggga ggccaaggaa ggtggatcac aaggtcagga gatcgagacc atcctggcta 2640 acacagtgaa accccgtctc tactaaaaat acaaaaaaat tagccaggct tggtggtggg 2700 cgcctatagt cccagctact tgggagactg aggcaggaga atgtcgtgaa cccgagaggc 2760 ggagcttgca gtgagccgag atcacgccac tgcactccag cctgggcgac agagtgacac 2820 tccgtctcaa aaaataaata tataaataaa taataaaaaa aaa 2863 <210> 23 <211> 1494 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 7474590CB1 <400> 23 atgtactctg acagcgagga tgagtcatca gagctcagca ctgtgctcag catgtttgag 60 gagaaggagt tcaccaggca gtacaccgtc ctgaagacct tgagccagca tggcactact 120 gaagtgaggc tatgctccca tcacctcaca ggtgtcacag ttgctgtcaa agctctgaag 180 taccagaggt ggtgggagcc aaaggtttca gaagtcgaaa tcatgaagat gctcagccac 240 cctaacattg tttcccttct gcaagtgata gagacagaac agaacattta tctgattatg 300 gaagtggccc aaggcacaca gctacataat cgagtccagg aggctaggtg cctgaaggaa 360 gatgaagcaa gaagcatatt tgttcagttg ctcagtgcca taggctactg tcatggtgaa 420 ggtgttgttc acagagacct aaagcctgac aatgtcatag ttgatgagca tggaaatgtc 480 aaaattgttg actttgggct aggtgccaga ttcatgcctg ggcagaaatt ggaaaggctg 540 tgtggagcct tccagttcat tcctccagag atattcctag ggctccctta tgatggccca 600 aaagtagaca tatgggcctt gggggttctt ttgtattata tggtgacagg gatttttcca 660 tttgtagggt ccaccttgtc agaaattagc aaggaagttc tacaagggag gtatgaaatt 720 ccttataatc tctctaaaga cttaaggagc atgataggcc tgttattggc aacaaacgca 780 aggcagaggc caactgcaca agacctccta agtcatccat ggcttcagga aggggaaaag 840 actatcacat ttcattccaa tggagacacc agctttccag accctgacat aatggcagcc 900 atgaaaaata ttgggtttca tgtgcaggac attagagaat cattaaaaca cagaaagttc 960 gatgaaacta tggctacata taacttactg agagctgagg catgtcagga tgatggcaat 1020 tatgttcaaa caaagttaat gaatccaggg atgccacctt tcccttcagt aacagactct 1080 ggagcttttt ctctgcctcc taggagaagg gccagtgaac cttcctttaa agtattagtc 1140 tcatctactg aagaacatca attaagacaa actgggggga caaatgcccc ttttccaccc 1200 aagaaaacac ccactatggg cagaagtcag aaacagaaac gtgccatgac tgccccttgt 1260 atttgtttac tgagaaacac ttacatagat acagaagaca gcagcttttg cactagctcc 1320 caggcagaaa agacttcaag tgatccagag aaaagtgaga cttcaacttc atgccctctg 1380 acacctaggg gctggaggaa atggaagaag agaattgtag catgcatcca gacattgtgt 1440 tgctgcacgt tgcctcaaaa aaaatgtccg aggagtgtgc atccccaaaa gtga 1494 <210> 24 <211> 2341 <212> DNA
<213> Homo sapiens <220>
<221> misc feature <223> Incyte ID No: 7474594CB1 <400> 24 atgtcagggc tggtgctgat gctggcggcg cggtgcattg tgggcagctc cccgctctgc 60 cgctgccgcc gccgtcgccc aaggaggatc ggggccgggc cgggccggga tgatccgggt 120 cggaaggccg ccgccgccgg agggagcggg tcacccaacg ccgcactgag ccgcccccgc i80 cccgccccgg ccccggggga tgcgccgccc cgagctgctg cctccgccgc cgccgcagcc 240 gcagccgcag cgggcacaga gcaggtagat ggccccctca gggcaggccc ggcggacacc 300 cctccctctg gctggcggat gcagtgccta gcggccgccc ttaaggacga aaccaacatg 360 agtgggggag gggagcaggc cgacatcctg ccggccaact acgtggtcaa ggatcgctgg 420 aaggtgctga aaaagatcgg gggcgggggc tttggtgaga tctacgaggc catggacctg 480 ctgaccaggg agaatgtggc cctcaaggtg gagtcagccc agcagcccaa gcaggtcctc 540 aagatggagg tggccgtgct caagaagttg caagggaagg accatgtgtg caggttcatt 600 ggctgtggca ggaacgagaa gtttaactat gtagtgatgc agctccaggg ccggaacctg 660 gccgacctgc gccgtagcca gccgcgaggc accttcacgc tgagcaccac attgcggctg 720 ggcaagcaga tcttggagtc catcgaggcc atccactctg tgggcttcct gcaccgtgac 780 atcaagcctt caaactttgc catgggcagg ctgccctcca cctacaggaa gtgctatatg 840 ctggacttcg ggctggcccg gcagtacacc aacaccacgg gggatgtgcg gccccctcgg 900 aatgtggccg ggtttcgagg aacggttcgc tatgcctcag tcaatgccca caagaaccgg 960 gagatgggcc gccacgacga cctgtggtcc ctcttctaca tgctggtgga gtttgcagtg 1020 ggccagctgc cctggaggaa gatcaaggac aaggaacagg tagggatgat caaggagaag 1080 tatgagcacc ggatgctgct gaagcacatg ccgtcagagt tccacctctt cctggaccac 1140 attgccagcc tcgactactt caccaagccc gactaccagt tgatcatgtc agtgtttgag 1200 aacagcatga aggagagggg cattgccgag aatgaggcct ttgactggga gaaggcaggc 1260 accgatgccc tcctgtccac gagcacctct accccgcccc agcagaacac ccggcagacg 1320 gcagccatgt ttggggtggt caatgtgacg ccagtgcctg gggacctgct ccgggagaac 1380 accgaggatg tgctacaggg agagcacctg agtgaccagg agaatgcacc cccaattctg 1440 cccgggaggc cctctgaggg gctgggcccc agtccccacc ttgtccccca ccccgggggt 1500 cctgaggctg aagtctggga ggagacagat gtcaaccgga acaaactccg gatcaacatc 1560 ggcaaagtaa ctgccgccag ggcgaagggc gtgggtggcc ttttctctca cccccgattc 1620 ccagccttgt gcccctgccc tgttcctcct aagcaccctg tccccggcca tctgcctgct 1680 tgccctgcct ctgtttcccg gtccctcccc gcactagcct cgctgtgtct tccatcatca 1740 tcatcctctg tctccttcac cctgaggaga ccatccgccc acagccgcct catcagcccc 1800 agctcatggc actcccctct cctgcagagc ccctgtgtgg aggaggaaca gagccgaggc 1860 atgggggtcc ccagctcccc agtgcgtgcc cccccagact cccccacaac cccagtccgt 1920 tctctgcgct accggagggt gaacagccct gagtcagaaa ggctgtccac ggcggacggg 1980 cgagtggagc tacctgagag gaggtgggtc tggggccagg ggcatggttg gggcccaagg 2040 ccctctccgc cttcacgtgg ctggtctgga ggaaaagtta gatgtgtggc ggaggtgggc 2100 agaccctggg aagtgctgag agggttatac ttgggcctgg ggtcagactc agttggggcc 2160 agagacaggg cctgggagaa ccagtggggg atccagagag gtcccggctc atgccaggaa 2220 acgtaattgg gtgagtgcag gctgcaggag ggacaggtgg ggcgcctggg cccaggaagg 2280 gtgaggggcg agttggttgg tgggtgtgtg tgcttccaga atctcttctc ctagagacta 2340 a 2341 <210> 25 <211> 2552 <212> DNA
<213> Homo Sapiens <220>
<221> misc feature <223> Incyte ID No: 7477585CB1 <400> 25 cgcggtgtct ggcgctcggt gggtgtggtt gcccctagtt tgaggcctgc ccgattaccc 60 gcaagacttg ggcagccccg ggcgccgctc cgaccacgac agggaaagga accttaatct 120 catctttaaa ataaggagaa ttactgagtg acctgaagga cccttttcag ctggaaagtc 180 tgaactgacc aacactggat gaatttgacc atttcttagg agactggaat gttaagtttc 240 tataaatgaa tgaaccagtt ctctcttgtt tggagcaatg ctgaaattcc aagaggcagc 300 taagtgtgtg agtggatcaa cagccatttc cacttatcca aagaccttga ttgcaagaag 360 atacgtgctt caacaaaaac ttggcagtgg aagttttgga actgtctatc tggtttcaga 420 caagaaagcc aaacgaggag aggaattaaa ggtacttaag gaaatatctg ttggagaact 480 aaatccaaat gaaactgtac aggccaattt ggaagcccaa ctcctctcca agctggacca 540 cccagccatt gtcaagttcc atgcaagttt tgtggagcaa gataatttct gcattatcac 600 ggagtactgt gagggccgag atctggacga taaaattcag gaatataaac aagctggaaa 660 aatctttcca gaaaatcaaa taatagaatg gtttatccag ctgctgctgg gagttgacta 720 catgcatgag aggaggatac ttcatcgaga cttaaagtca aagaatgtat ttctgaaaaa 780 taatctcctt aaaattggag attttggagt ttctcgactt ctaatgggat cctgtgacct 840 ggccacaact ttaactggaa ctccccatta tatgagtcct gaggctctga aacaccaagg 900 ctatgacaca aagtcggaca tctggtcact ggcatgcatt ttgtatgaga tgtgctgcat 960 gaatcatgca ttcgctggct ccaatttctt atccattgtt ttaaaaattg ttgaaggtga 1020 cacaccttct ctccctgaga gatatccaaa agaactaaat gccatcatgg aaagcatgtt 1080 gaacaagaat ccttcattaa gaccatctgc tatcgaaatt ttaaaaatcc cttaccttga 1140 tgagcagcta cagaacctaa tgtgtagata ttcagaaatg actctggaag acaaaaattt 1200 ggattgtcag aaggaggctg ctcatataat taatgccatg caaaaaagga tccacctgca 1260 gactctgagg gcactgtcag aagtacagaa aatgacgcca agagaaagga tgcggctgag 1320 gaagctccag gcggctgatg agaaagccag gaagctgaaa aagattgtgg aagaaaaata 1380 tgaagaaaat agcaaacgaa tgcaagaatt gagatctcgg aactttcagc agctgagtgt 1440 tgatgtactc catgaaaaaa cacatttaaa aggaatggaa gaaaaggagg agcaacctga 1500 gggaagactt tcttgttcac cccaggacga ggatgaagag aggtggcaag gcagggaaga 1560 ggaatctgat gaaccaactt tagagaacct gcctgagtct cagcctattc cttccatgga 1620 cctccacgaa cttgaatcaa ttgtagagga tgccacatct gaccttggat accatgagat 1680 cccagaagac ccacttgtgg ctgaagagta ctacgctgat gcatttgatt cctattgtgt 1740 agagagtgat gaggaggaag aagaaatagc gttagaaaga ccagagaaag aaatcaggaa 1800 tgagggatcc cagcctgctt acagaacaaa ccaacaggac agtgatatcg aagcgttggc 1860 caggtgtttg gaaaatgtcc tgggttgcac ttctctagac acaaagacca tcaccaccat 1920 ggctgaagac atgtccccag gaccaccaat tttcaacagt gtgatggcca ggaccaagat 1980 gaaacgcatg agggaatcag ccatgcagaa gctggggaca gaagtatttg aagaggtcta 2040 taattacctc aagagagcaa ggcatcagaa tgctagcgaa gcagagatcc gcgagtgttt 2].00 ggaaaaagtg gtgcctcaag ccagcgactg ttttgaagtg gaccagctcc tgtactttga 2160 agagcagttg ctgatcacga tgggaaaaga acctactctc cagaaccatc tctaggcaac 2220 tatcaaaaag aagcagaagt tcaagtggac aaatttatgt gaaaattcat ttaacatata 2280 agctgaactc tattatgggg aatggataca aaagcagagc tcccatcttg actttcaatt 2340 cctcatcaga agtactggct tctttagaga gtagtaagca tggctgccta tgcttggagt 2400 cataagtgtt atttggacta taccctgaga taagcttata gatcaagttt ggctcccttg 2460 aaaagcattt ctctcatgtg cgccctcagg gcttccagca ggattgagtc accctgacga 2520 tgaccgggga gaagccgtgt gctcttcatt at 2552 <210> 26 <211> 2176 <212> DNA
<213> Homo sapiens <220>
<221> misc feature <223> Incyte ID No: 7477587CB1 <400> 26 ctcaccgccc tccccaccag ccccagcatc acaaacgctg ggcttctctg ccgctccgaa 60 ttctgctgtg gctccccagt gcccagggcc atcaagcccc aggttttcat gtgggcccca 120 gggacccgcc agcagggagg acccgaaatg gcacacattc agaacgtcga ggctcatacc 180 tcaagcgcac tgtgggggag aagcccccgg aagcccccaa ccccccacgc gcgagagagc 240 ctcagtttcc cgctcgagcg gccccggagc ggccgcagcg cggtggtctc ggcccggctg 300 cgccagagtc cgcgcatgga gccccggccg cggcggcggc gcaggagtcg ccccctggte 360 gccgccttcc tgcgagaccc gggctcgggc cgcgtgtaca ggcgcgggaa gctgatcggc 420 aagggcgcct tcagccgctg ctacaagctg acagacatgt ccaccagcgc cgtgttcgcc 480 ctcaaggtgg tgccgtgtgg cggggctggg gccgggtggc ttcgcccgca gggaaaggtg 540 gagcgtgaga ttgccctgca tagccgcctg cgaccccgca acatcgtggc tttccacgga 600 cactttgctg accgcgacca cgtgtacatg gtgctggagt actgcagccg ccagtctttg 660 gcccacgtgc tgagggcgcg gcagatcctg acggagccag aagtgcgcga ctacctgcgg 720 ggcctggtca gcggcctgcg ctacctgcac cagcggtgca tcctgcaccg cgacctgaag 780 ctcagtaact tcttccttaa caagaacatg gaggtgaaga ttggagacct gggactggcg 840 gccaaggtgg ggccaggggg ccgctgccac aggtacacgg tgctgactgg caccccaccc 900 ttcatggcct cacccctgtc ggagatgtac caaaacatcc gtgagggcca ctaccccgaa 960 cccgctcacc tgtctgccaa tgcgcgccgc ctcatcgtgc acctcctagc acccaacccg 1020 gccgagcggc ccagcctgga ccacctgctg caggacgact tcttcacaca gggtttcact 1080 ccagaccggc tgccggccca ctcctgccac agtcccccca tcttcgccat acccccgcct 1140 ctgggcagga tcttccggaa ggtgggccag cggctgctca cccagtgccg gccaccctgc 1200 cccttcacgc ctaaagaggc ctcgggtcca ggagaaggtg ggccagaccc tgactccatg 1260 gagtgggacg gcgagagctc cctgtctgcg aaagaggttc cctgcctgga aggccccatc 1320 cacctggtcg cacaagggac cctgcagagt gacctggccg ccacacagga ccccctggga 1380 gagcagcagc ccatcctctg ggcccccaaa tgggtggatt attccagcaa atacggcttt 1440 ggctaccagc tcttggacgg ggggcgcacg ggacggcacc cacatggccc tgcgaccccc 1500 cggaggtatt tattaagcac ctactgtgca cacctacagg tgctccctgc ctgccaagtg 1560 tgctacatgc ccaactgcgg gaggctggaa gccttcgccc tgagggatgt gcccggcctg 1620 ctgggcgcca agctggccgt gctgcagctc tttgccggct gcctgcggcg gcggctgcgg 1680 gaggagggga ccctccccac acctgtgcca cctgctggac ccggcctctg cctcctgcgc 1740 ttcctggcct ctgagcacgc cctgctgctg ctgttcagca atgggatggt gcaggtgagc 1800 ttcagtggag tcccggccca actggtgctg agtggcgagg gtgagggttt gcagctcacc 1860 ctctgggagc aggggtcccc tggcacctcc tactccctgg acgtcccgcg gagccacggc 1920 tgcgccccca ccaccggaca gcaccttcac cacgccctcc gcatgctgca gagtatctag 1980 tgcccctgag ggtcagagtg gacccctgca tggtagtgcc agggacccag gctccatttc 2040 cattcctgtg gctcccccag aggggctgtc ctgggggaga gctggggggc acacgggagg 2100 tgggttcttg ccttgtggca tgactgttca acccagactt tgctgggatc tcttcctttt 2160 tcattaaaga caattc 2176 <210> 27 <211> 4277 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 7594537CB1 <400> 27 tgtgagcaga gttcttgaag ctccactcct cctggggaag ccgagctgtg tgggagcctt 60 cttactgtgc cgggagcgtg tgaattggaa aggatcctga gaactggcta gtcccagttc 120 ctctccggaa aagcagtggc tctcgcttca gagatgcgct cagctttcgc ctgcatcaca 180 ctgcattctt taacacatta ttgaaattac aagcatccaa agcagtttca tgtggacaga 240 ttgcatattt tgaaagcctg aggtatttta tcatgaaaca tgccatgtgg aatctttgaa 300 gcatagacct ctgcgcaaca cctgaataaa gaatctttta cctggtatgt gacagagctt 360 ctcaccacca ccatgacaaa ccaggaaaaa tgggcccacc tcagcccttc ggaattttcc 420 caacttcaga aatatgctga gtattctaca aagaaattaa aggatgttct tgaagaattc 480 catggtaatg gtgtgcttgc aaagtataat cctgaaggga caatagattt tgaaggtttc 540 aaactattca tgaagacatt cctggaagcc gagcttcctg atgatttcac tgcacacctt 600 ttcatgtcat ttagcaacaa gtttcctcat tctagtccaa tggtaaaaag taagcctgct 660 ctcctatcag gcggtctgag aatgaataaa ggtgccatca cccctccccg aactacttct 720 cctgcaaata cgtgttcccc agaagtaatc catctgaagg acattgtctg ttacctgtct 780 ctgcttgaaa gaggaagacc tgaggataag cttgagttta tgtttcgcct ttatgacacg 840, gatgggaatg gcttcctgga cagctcggag ctagaaaata tcatcagtca gatgatgcat 900 gttgcagaat accttgagtg ggatgtcact gaacttaatc caatcctcca tgaaatgatg 960 gaagaaattg actatgatca tgatggaacc gtgtctctgg aggaatggat tcaaggagga 1020 atgacaacga ttccacttct tgtgetcctg ggcttagaaa ataacgtgaa ggatgatgga 1080 cagcacgtgt ggcgactgaa gcactttaac aaacctgcct attgcaacct ttgcctgaac 1140 atgctgattg gcgtggggaa gcagggcctc tgctgttcct tctgcaagta cacagtccat 1200 gagcgctgtg tggctcgagc acctccctct tgcatcaaga cctatgtgaa gtccaaaagg 1260 aacactgatg tcatgcacca ttactgggtt gaaggtaact gcccaaccaa gtgtgataag 1320 tgccacaaaa ctgttaaatg ttaccagggc ctgacaggac tgcattgtgt ttggtgtcag 1380 atcacactgc ataataaatg tgcttctcat ctaaaacctg aatgtgactg tggacctttg 1440 aaggaccata ttttaccacc cacaacaatc tgtccagtgg tactgcagac tctgcccact 1500 tcaggagttt cagttcctga ggaaagacaa tcaacagtga aaaaggaaaa gagtggttcc 1560 cagcagccaa acaaagtgat tgacaagaat aaaatgcaaa gagccaactc tgttactgta 1620 gatggacaag gcctgcaggt cactcctgtg cctggtactc acccactttt agtttttgtg 1680 aaccccaaaa gtggtggaaa acaaggagaa cgaatttaca gaaaattcca gtatctatta 1740 aatcctcgtc aggtttacag tctttctgga aatggaccaa tgccagggtt aaactttttc 1800 cgtgatgttc ctgacttcag agtgttagcc tgtggtggag atggaaccgt gggctgggtt 1860 ttggattgca tagaaaaggc caatgtaggc aagcatcctc cagttgcgat tctgcctctt 1920 gggactggca atgatctagc aagatgcctg cgatggggag gaggttacga aggtgagaat 1980 ctgatgaaaa ttctaaaaga cattgaaaac agcacagaaa tcatgttgga caggtggaag 2040 tttgaagtca tacctaatga caaagatgag aaaggagacc cagtgcctta cagtatcatc 2100 aataattact tttccattgg cgtggatgcc tccattgcac acagattcca catcatgaga 2160 gaaaaacacc cagagaaatt caacagtaga atgaagaaca aattttggta ttttgagttt 2220 ggcacatctg aaactttctc agccacctgc aagaagctac atgaatctgt agaaatagaa 2280 tgtgatggag tacagataga tttaataaac atctctctgg aaggaattgc tattttgaat 2340 ataccaagca tgcatggagg atccaatctt tggggagagt ctaagaaaag acgaagccat 2400 cgacgaatag agaaaaaagg gtctgacaaa aggaccaccg tcacagatgc caaagagttg 2460 aagtttgcaa gtcaagatct cagtgaccag ctgctggagg tggtcggctt ggaaggagcc 2520 atggagatgg ggcaaatata cacaggcctg aaaagtgctg gccggcggct ggctcagtgc 2580 tcctgcgtgg tcatcaggac gagcaagtct ctgccaatgc aaattgatgg ggagccatgg 2640 atgcagaccc catgcacaat aaaaattaca cacaagaacc aagccccaat gctgatgggc 2700 ccgcctccaa aaaccggttt attctgctcc ctcgtcaaaa ggacaagaaa ccgaagcaag 2760 gaataatcct gtgttgtttc actcttagaa attgaattag cataattggg ccatggaaca 2820 catatgctgg aaatctttga accatttcaa gtctcctgct catgcaaaat catggaagtg 2880 gtttaacagt ttttgttact aagctaatgt aaaattcagc tattagaaaa tttattgtct 2940 cagtttttat aggcatcttt gcatgaagaa agcagaagtt tacctgaagt gatactgcat 3000 atttttggtg catgcattcc catagatttt tacatctccc acccaactct tccccaattt 3060 ccttttacta acctgtgaga aaaacccgtg aaacatgaaa aaggaaatac catgggaaac 3120 gtgattctca gtgtgattcc aattattacg aagcactaat cagtaacgct acaatgatca 3180 taattgcaga ttgctatacg tttccctttt agaatcagtg tatcagtgac ctatgacttg 3240 aggagaaact tttaattcga agattttatt aaatagttga ctacaatacc ttgctatata 3300 tacatagttt ttcttcaaca tcttaactct tctgagtgga aataaaaata tcaggcataa 3360 ggttttctca tgctgaaaaa tagaacgcgg tttttatttt gcttagtttt ctttttaatt 3420 ccagaaataa gtgaaaacat gttacttgac agtcaagtgt ggtaatatgg caagccttgt 3480 tcctttctgc atgagaatct aggagagaat tcataaccac accaataacg aaatagaagt 3540 tttaaactat gtgcctaatc aatgtgtttc ccaccaaaga ttcagaaaac aatgcttgag 3600 agaaatgggt taatgcataa ttaattaagc attgtggagc aaatttaggg ttcctgtgat 3660 taattttgtg atgactaaaa tgctggaaag caagtgagtt gcccattaat tatgattaaa 3720 attctcacct ttcacagaca gacaataagc cagacaacac aatcaaagct caatagatga 3780 tttcttgctt ttttcagtca tttataaata taggtgtaat ttttcatgga tcagttaagt 3840 acacttgaag gaagtaaatg attgtatcag tttatttcta gtataaatgg gtacctgtaa 3900 taatactgag ctcttggaag cgaatcatgc atgcaattag ctccctcctc ctcacctact 3960 ccactcccat ctttatgaca tttcaaatgt ttatttggaa acaacagcct agatcactgt 4020 tgaaggtgtt catggcatag ttggagtctc tgactgttta aagaaatcac agaacagtac 4080 ttttctttta gtgtttcatt aagcctatga tgtaaaatga aatgcttctg agcagtcttg 4140 taatattgtt cattcatatt gacctgcatc tcatcattgc atgttttatg ttttcaaaca 4200 tgccataagg aaaacgagtg cctgaactgc atgatttatt agtttctctc cactctgcat 4260 taaagtgcta atgattt 4277 <210> 28 <211> 2616 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 70467491CB1 <400> 28 atgtccacta ggaccccatt gccaacggtg aatgaacgag acactgaaaa cgctgtattg 60 ccgcacacgt cacatggaga tgggcgtcaa gaagttacct ctcgtaccag ccgctcagga 120 gctcggtgta gaaactctat agcctcctgt gcagatgaac aacctcacat cggaaactac 180 agactgttga aaacaatcgg caaggggaat tttgcaaaag taaaattggc aagacatatc 240 cttacaggca gagagaaaaa tgttagaata tccaaagaaa ttgataattt tctaggaaaa 300 catgacttac caaaattaac tctagaaaag aatcgataca catcagtaac aacagaagtt 360 gagaaagtag ttaacatatt gccaaacctg gaattcatga ttgaattctt tgagatctac 420 tctataggtg aagtatttga ctatttggtt gcacatggca ggatgaagga aaaagaagca 480 agatctaaat ttagacagat tgtgtctgca gttcaatact gccatcagaa acggatcgta 540 catcgagacc tcaaggctga aaatctattg ttagatgccg atatgaacat taaaatagca 600 gatttcggtt ttagcaatga atttactgtt ggcggtaaac tcgacacgtt ttgtggcagt 660 cctccatacg cagcacctga gctcttccag ggcaagaaat atgacgggcc agaagtggat 720 gtgtggagtc tgggggtcat tttatacaca ctagtcagtg gctcacttcc ctttgatggg 780 caaaacctaa aggaactgag agagagagta ttaagaggga aatacagaat tcccttctac 840 atgtctacag actgtgaaaa ccttctcaaa cgtttcctgg tgctaaatcc aattaaacgc 900 ggcactctag agcaaatcat gaaggacagg tggatcaatg cagggcatga agaagatgaa 960 ctcaaaccat ttgttgaacc agagctagac atctcagacc aaaaaagaat agatattatg 1020 gtgggaatgg gatattcaca agaagaaatt caagaatctc ttagtaagat gaaatacgat 1080 gaaatcacag ctacatattt gttattgggg agaaaatctt cagagctgga tgctagtgat 1140 tccagttcta gcagcaatct ttcacttgct aaggttaggc cgagcagtga tctcaacaac 1200 agtactggcc agtctcctca ccacaaagtg cagagaagtg tttcttcaag ccaaaagcaa 1260 agacgctaca gtgaccatgc tggaccagct attccttctg ttgtggcgta tccgaaaagg 1320 agtcagacca gcactgcaga tagtgacctc aaagaagatg gaatttcctc ccggaaatca 1380 agtggcagtg ctgttggagg aaagggaatt gctccagcca gtcccatgct tgggaatgca 1440 agtaatccta ataaggcgga tattcctgaa cgcaagaaaa gctccactgt ccctagtagt 1500 aacacagcat ctggtggaat gacacgacga aatacttatg tttgcagtga gagaactaca 1560 gctgatagac actcagtgat tcagaatggc aaagaaaaca gcactattcc tgatcagaga 1620 actccagttg cttcaacaca cagtatcagt agtgcagcca ccccagatcg aatccgcttc 1680 ccaagaggca ctgccagtcg tagcactttc cacggccagc cccgggaacg gcgaaccgca 1740 acatataatg gccctcctgc ctctcccagc ctgtcccatg aagccacacc attgtcccag 1800 actcgaagcc gaggctccac taatctcttt agtaaattaa cttcaaaact cacaaggagg 1860 cttccaactg aatatgagag gaacgggaga tatgagggct caagtcgcaa tgtatctgct 1920 gagcaaaaag atgaaaacaa agaagcaaag cctcgatccc tacgcttcac ctggagcatg 1980 aaaaccacta gttcaatgga tcccggggac atgatgcggg aaatccgcaa agtgttggac 2040 gccaataact gcgactatga gcagagggag cgcttcttgc tcttctgcgt ccacggagat 2100 gggcacgcgg agaacctcgt gcagtgggaa atggaagtgt gcaagctgcc aagactgtct 2160 ctgaacgggg tccggtttaa gcggatatcg gggacatcca tagccttcaa aaatattgct 2220 tccaaaattg ccaatgagct aaagctgtaa cccagtgatt atgatgtaaa ttaagtagca 2280 attaaagtgt tttcctgaac actgatggaa atgtatagaa taatatttag gcaataacgt 2340 ctgcatcttc taaatcatga aattaaagtc tgaggacgag agcaaaaaaa aaaaaaaagg 2400 gcggccctcg agccgctcga gccgaattcg gctcgaggat tcagtgggtg agagggaaga 2460 aggggaggtt ggggggctcc ttcccttcag aacttgaagt ttctcccact gcctcctctc 2520 cagtggtctc ccaggtgcca gacccaaaag cttttcctac agtgataccc ttatattttt 2580 acttcccctt gactcatatg ttttaacatg aatttt 2616 <210> 29 <211> 1253 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 7478559CB1 <400> 29 ctggcccctc ctctaccact cccactccct cgccggaccc ccccgccggg gctagcgtct 60 gccgcggctc cgagggggtg gggctgctgg gaatggctgt gcccccttcg gcccctcagc 120 cgcgcgcgtc ctttcacctg aggaggcaca cgccttgccc gcagtgctca tggggcatgg 180 aggagaaggc ggcggccagc gccagctgcc gggagccgcc gggccccccg agggccgccg 240 ccgtcgcgta cttcggcatt tccgtggacc cggacgacat ccttcccggg gccctgcgcc 300 tcatccagga gctgcggccg cattggaaac ccgagcaagt tcggaccaag cgcttcatgg 360 atggcatcac caacaagctg gtggcctgct atgtggagga ggacatgcag gactgcgtgc 420 tggtccgggt gtatggggag cggacggagc tgctggtgga ccgggagaat gaggtcagaa 480 acttccagct gctgcgagca cacagctgtg cccccaaact ctactgcacc ttccagaatg 540 ggctgtgcta tgagtacatg cagggtgtgg ccctggagcc tgagcacatc cgtgagcccc 600 ggcttttcag gttaatcgcc ttagaaatgg caaagattca tactatccac gccaacggca 660 gcctgcccaa gcccatcctc tggcacaaga tgcacaatta tttcacgctt gtgaagaacg 720 agatcaaccc cagcctttct gcagatgtcc ctaaggtaga ggtgttggaa cgggagctgg 780 cctggctgaa ggagcatctg tcccagctgg agtcccctgt ggtgttttgt cacaatgacc 840 tgctctgcaa gaatatcatc tatgacagca,tcaaaggtca cgtgcggttc attgactatg 900 aatatgctgg ctacaactac caagcttttg acattggcaa ccatttcaat gagtttgcag 960 gcgtgaatga ggtggattac tgcctgtacc cggcgcggga gacccagctg cagtggctgc 1020 actactacct gcaggcacaa aaggggatgg ccgtgacccc cagggaggtg caaaggctct 1080 acgtgcaagt caacaagttt gccctggcgt ctcacttctt ctgggctctc tgggccctca 1140 tccagaacca gtactccacc atcgactttg atttcctcag gtacgcagtg atccgattca 1200 accagtactt caaggtgaag cctcaagcgt cagccttgga gatgccaaag tga 1253 <210> 30 <211> 1790 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1698381CB1 <400> 30 tttaagaagg agttccctta taggagatgg aagaaacggc cattaatccg gggacttttt 60 atgctggaaa caaacctgaa ggtacaggtt cggcccggaa gttataccca ccaagagaag 120 tatgtccgga attgtgggtt ctgcagtcac tgacttcaag aatgaagccg cggaccctcg 180 cggtgcagca ttgtactgca agtcaatcga tacaataatt taagtcactt cagctataat 240 ggaaaagtat gaaaaattag ctaagactgg agaagggtct tatggggttg tattcaaatg 300 cagaaacaaa acctctggac aagtagtagc tgttaaaaaa tttgtggaat ctgaagatga 360 tcctgttgtt aagaaaatag cactaagaga aatacgtatg ttgaagcaat taaaacatcc 420 aaatcttgtg aacctcatcg aggtgttcag gagaaaaagg aaaatgcatt tagtttttga 480 atactgtgat catacacttt taaatgagct ggaaagaaac ccaaatggag ttgctgatgg 540 agtgatcaaa agcgtattat ggcaaacact tcaagctctt aatttctgtc atatacataa 600 ctgtattcac agagatataa aacctgaaaa tattctaata actaagcaag gaataatcaa 660 gatttgtgac ttcgggtttg cacaaattct gattccagga gatgcctaca ccgattatgt 720 agctacgaga tggtaccgag ctcctgaact tcttgtggga gatactcagt atggttcttc 780 agtcgatata tgggctattg gttgtgtttt tgcagagctc ctgacaggcc agccactgtg 840 gcctggaaaa tcagatgtgg accaacttta tctgataatc agaacactag gaaaattaat 900 cccaagacat caatcaatct ttaaaagtaa cgggtttttc catggcatca gtatacctga 960 gccagaagac atggaaactc ttgaggaaaa gttctcagat gttcatcctg tggctctgaa 1020 cttcatgaag gggtgtctga agatgaatcc agatgacaga ttaacctgtt cccaactcct 1080 ggagagctcc tactttgatt cttttcaaga ggcccaaatt aaaagaaaag cacgtaatga 1140 aggaagaaac agaagacgcc aacagaatca actgttgcct ctcataccag gaagccacat 1200 ctcccccaca cctgatggaa gaaaacaagt cctccagtta aaatttgatc accttccaaa 1260 catttaggaa aatgttcttt caagtgcaaa gtaatttaat atgtacacat tttgtacaag 1320 tgagatagga attctcagtg tttcaaatgc aaatgagcca tatgaaaatt aagatgcctt 1380 ctagaattgt tttggctctg atcattgctg attcctttcc ccatgctttt acatgccaac 1440 tttatctttt agaatatttt ctttaaatgt tataaagcct aaaactgcac atatggaaga 1500 gacattttca atttcatcag agcagcccct cccgaggcta tctatatgga gaatttgtga 1560 gcttatactt ggatttatga aaaagattta catgtgtcat cttgcttcag ctgaccacat 1620 aatttcttaa agcaatatca aatagcctgc ctcactgttt gtgtaagaaa tgacatatgt 1680 tcctgcatgt gtaattcata cttattgtaa ccaggtctgt tgagtattgc tggtatctta 1740 tactgagtaa atatggtgta gaaagggaac tttgaagggc tgcagattcg 1790 <210> 31 <211> 4132 <212> DNA
<213> Homo Sapiens <220>
<221> misc feature <223> Incyte ID No: 7474637CB1 <400> 31 cccccgactg tcttggtggc agaggggact tttattcagc tggaaccgcg cggcgaggcc 60 caagtgtctc tggagagatt cggggttcag gaggtggcgg gtgcacccaa gggtgctggg 120 aggaagctcc aggttcccat tcttccccag ggatcggcgt tgcccctgct cgcgggggta 180 gtctagggca acggaagatg gcggcggcgg ccgggcacgg ggttccgggc tccgctcggg 240 cagagcccac ccgctgacca actccgccgc ccccgccggg cggtgctgtg tccccgcagg 300 agtcggagag gatggcaggg gccggaggcc agcaccaccc tccgggcgcc gctggaggag 360 cggccgccgg agccggcgcc gcggtcacct ccgccgctgc ctcggcgggg ccgggagagg 420 attcgtctga cagcgaagcg gagcaagagg gaccccagaa actgatccgc aaagtgtcta 480 cctcggggca gatccggacc aagaccagta ttaaagaggg acagctattg aagcaaacca 540 gttctttcca aaggtggaaa aagcgatact tcaaacttcg aggccgcacc ctttactatg 600 caaaggactc aaagtctctg atatttgatg aagttgacct ctcagatgct agtgtagctg 660 aagcaagcac gaaaaatgct aacaacagct tcacgatcat cactccattc agaaggctaa 720 tgctgtgtgc tgagaacaga aaggagatgg aggattggat cagctcactg aagtctgtac 780 agaccagaga accctacgag gtggcccagt ttaatgtgga acatttctca gggatgcaca 840 actggtacgc ctgctcccac gcccgaccca ccttctgtaa cgtgtgcaga gagagtcttt 900 ctggagtcac ctcccatggc ctgtcctgcg aagtgtgtaa attcaaggct cacaaaagat 960 gtgcagtgag agcaacaaat aactgtaaat ggactaccct ggcctccatc gggaaggaca 1020 ttatagaaga tgaagatggc gtcgcgatgc ctcaccagtg gcttgagggc aacctgcctg 1080 taagtgccaa gtgtgctgtc tgcgacaaaa catgtggcag tgttctccgt ctacaggatt 1140 ggaaatgcct ttggtgtaag acaatggtac acactgcctg caaagattta taccatccaa 1200 tatgtccact tggtcaatgt aaagtatcta tcatacctcc aattgcacta aacagcaccg 1260 attccgatgg tttctgtaga gcaacatttt cgttctgtgt tagtcctcta ttggtttttg 1320 tcaattctaa gagtggagat aatcagggag taaagttcct ccgtcgcttt aaacagttgc 1380 taaatccggc tcaggtgttt gatttaatga atggaggtcc tcatttaggt ttaagattat 1440 ttcagaagtt tgacaatttc cggattcttg tatgtggagg cgatggaagt gtaggttggg 1500 ttttgtcaga aatcgataag ctcaacttga ataaacagtg tcagctggga gtgttgcctt 1560 tgggtacagg aaatgacctt gcccgagttc ttggctgggg aggttcatat gacgatgaca 1620 cccagcttcc tcagatccta gagaaactgg aacgagccag taccaaaatg ttggacaggt 1680 ggagtataat gacatatgaa ctcaaattgc caccaaaagc ttccctactt ccaggacctc 1740 cagaagcatc tgaagaattt tatatgacga tttatgaaga ctcagttgca acgcatctta 1800 caaaaatcct caattctgat gaacatgcag tggtcatatc ttctgccaag acgctatgtg 1860 aaactgtaaa ggacttcgtt gccaaagtag aaaagacgta tgacaaaacc ttggaaaatg 1920 ccgttgtagc tgatgccgtg gccagtaaat gttcagtcct aaacgagaag ctcgaacaac 1980 tgctgcaggc tttgcacaca gattcccagg ctgcgcctgt tctccctggc ctcagccctc 2040 tcattgtgga agaagatgct gtggaatcgt ccagtgaaga gtccctgggt gaaagcaaag 2100 agcagcttgg ggatgacgtt acaaaacctt cctcccagaa agccgtcaaa ccaagggaaa 2160 tcatgttgcg ggcaaatagt ttaaagaaag cagtgaggca agtcattgag gaagccggaa 2220 aagttatgga tgacccgaca gttcacccct gtgaaccagc taatcagtcc tctgattatg 2280 acagcacaga aacagatgaa tctaaggagg aagctaaaga tgatggtgcc aaagaatcaa 2340 taactgttaa aactgcacct cggtctccag atgcccgggc aagttatggc cattcccaaa 2400 ctgattctgt ccctggtcca gctgtggcag ccagcaaaga aaacctccct gtgctcaata 2460 ccagaataat ctgcccaggt ttaagagcag gactggctgc ctcaattgct gggagttcga 2520 ttatcaacaa aatgttactg gcaaacattg atccttttgg tgccacgccg tttattgacc 2580 ctgatctaga ttccgtagat ggatattcag aaaaatgtgt catgaacaat tactttggga 2640 ttggattaga tgcaaaaatt tcattagaat ttaataataa aagagaggag caccctgaaa 2700 aatgcaggag ccgaactaaa aacttgatgt ggtatggagt ccttggaacc cgggagttat 2760 tacagagatc gtacaagaat ttagaacaaa gggttcaact tgagtgtgat gggcagtata 2820 ttcctcttcc cagcttgcaa ggcatagccg tgttgaacat tcccagctat gctggaggca 2880 ctaacttttg gggtggaact aaagaggatg atatatttgc tgcaccatcc tttgatgaca 2940 agatcctgga agttgtagca atatttgata gcatgcaaat ggcagtttca agggtcatta 3000 aactgcagca tcatcgaata gcccagtgcc gtacagtgaa aatcactata tttggtgacg 300'0 aaggagtccc agtgcaagtg gatggtgaag cgtgggttca gcctccaggg attatcaaaa 3120 ttgtgcacaa aaacagagca caaatgctaa caagggacag agcctttgag agcactctga 3180 aatcttggga agataagcag aagtgtgatt ctggtaaacc agttctccga acccatttgt 3240 acatccatca cgccattgac ttggcaacag aagaggtgtc gcagatgcag ctatgctccc 3300 aggctgcaga ggagctcatt actaggatat gtgacgcagc cacaattcac tgtcttttgg 3360 agcaagaact ggcccatgct gtgaatgcct gctcccatgc cctgaataaa gccaacccaa 3420 ggtgcccgga gagtcttaca agagacactg ccactgaaat agccatcaat gtgaaggcgc 3480 tgtataatga aacagaatct ttgctagttg gcagggttcc tttgcagctg gaatcgccac 3540 atgaagagcg agtatccaat gccttacact ctgtggaggt ggaattacag aaactgacag 3600 agattccttg gctttattat atcttacacc caaatgaaga tgaggaacct cctatggatt 3660 gcaccaaaag gaacaacaga agcaccgtat ttcgaatagt gccaaagttt aaaaaggaaa 3720 aggttcagaa gcagaagaca agttcacagc ctggatctgg ggataccgaa agtgggtcat 3780 gtgaagcgaa ttctccaggg aattaaagag cttggaagga gcactccaca gtcggaggtg 3840 taatcatatt ggtgctattc cttggaagag aagttattgc cacttaatac aaagtccttg 3900 gaagcaagtg gctgttcttg tagttttctg catagataag taagcaccac tgaagcacct 3960 ctgtggcttg atattttgct gtgggtgaaa ttttgatttg aggtattaga aaatattttt 4020 gtgccgaaca atacattcca cgaagccatt ttctttttgt gcaaacctga catgttcaaa 4080 tatattcaca atggtaataa ggtaggagga atctgagacg attgcattgt ct 4132 <210> 32 <211> 1137 <212> DNA
<213> Homo sapiens <220>
<221> misc feature <223> Incyte ID No: 7170260CB1 <400> 32 atggaggact ttctgctctc caatgggtac cagctgggca agaccattgg ggaagggacc 60 tactcaaaag tcaaagaagc attttccaaa aaacaccaaa gaaaagtggc aattaaagtt 120 atagacaaga tgggagggcc agaagagttt atccagagat tcctccctcg ggagctccaa 180 atcgtccgta ccctggacca caagaacatc atccaggtgt atgagatgct ggagtctgcc 240 gacgggaaaa tctgcctggt gatggagctc gctgagggag gggatgtctt tgactgcgtg 300 ctgaatgggg ggccactgcc tgaaagccgg gccaaggccc tcttccgtca gatggttgag 360 gccatccgct actgccatgg ctgtggtgtg gcccaccggg acctcaaatg tgagaacgcc 420 ttgttgcagg gcttcaacct gaagctgact gactttggct ttgccaaggt gttgcccaag 480 tcacaccggg agctgagcca gaccttctgc ggcagtacag cctatgctgc ccccgaggtg 540 ctgcagggca ttccccacga tagcaaaaaa ggtgatgtct ggagcatggg tgtggtcctg 600 tatgtcatgc tctgtgccag cctacctttt gacgacacag acatccccaa gatgctgtgg 660 cagcagcaga agggggtgtc cttccccact catctgagca tctcggccga ttgccaggac 720 ctgctcaaga ggctcctgga acccgatatg atcctccggc cttcaattga agaagttagt 780 tggcatccat ggctagcaag cacttgataa aagcaatggc aagtgctctc caataaagta 840 gggggagaaa gcaaacccaa aaacccgctt ctaaaatggt gatatatatt ttacgcttta 900 agtttactta tcctaaaact tacctacatc taccccagcc ttactactac tctttccttt 960 tagagatctt catggaatca aagggcctca ttcagacttc cttttttttt ttaagagtct 1020 tgctctgtcg cccaggctgg aatgcagtgg cacgattcca gttcactgca actctgcttc 1080 ccaggttcaa gcgattctcc tgcctcagcc tccccagtag ctggctttcc agcacac 1137 <210> 33 <211> 3365 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1797506CB1 <400> 33 atgagaaggg cggggatcgg cgaggactcc aggctggggt tgcaggccca gccaggggcg 60 gagccttctc cgggtcgggc ggggacagag cgctcccttg gaggcaccca gggacctggc 120 cagccgtgca gctgcccagg cgctatggcg agtgcggtca gggggtcgag gccgtggccc 180 cggctggggc tccagctcca gttcgcggcg ctgctgctcg ggacgctgag tccacaggtt 240 catactctca ggccagagaa cctcctgctg gtgtccacct tggatggaag tctccacgca 300 ctaagcaagc agacagggga cctgaagtgg actctgaggg atgatcccgt catcgaagga 360 ccaatgtacg tcacagaaat ggcctttctc tctgacccag cagatggcag cctgtacatc 420 ttggggaccc aaaaacaaca gggattaatg aaactgccat tcaccatccc tgagctggtt 480 catgcctctc cctgccgcag ctctgatggg gtcttctaca caggccggaa gcaggatgcc 540 tggtttgtgg tggaccctga gtcaggggag acccagatga cactgaccac agagggtccc 600 tccacccccc gcctctacat tggccgaaca cagtatacgg tcaccatgca tgacccaaga 660 gccccagccc tgcgctggaa caccacctac cgccgctact cagcgccccc catggatggc 720 tcacctggga aatacatgag ccacctggcg tcctgcggga tgggcctgct gctcactgtg 780 gacccaggaa gcgggacggt gctgtggaca caggacctgg gcgtgcctgt gatgggcgtc 840 tacacctggc accaggacgg cctgcgccag ctgccgcatc tcacgctggc tcgagacact 900 ctgcatttcc tcgccctccg ctggggccac atccgactgc ctgcctcagg cccccgggac 960 acagccaccc tcttctctac cttggacacc cagctgctaa tgacgctgta tgtggggaag 1020 gatgaaactg gcttctatgt ctctaaagca ctggtccaca caggagtggc cctggtgcct 1080 cgtggactga ccctggcccc cgcagatggc cccaccacag atgaggtgac actccaagtc 1140 tcaggagagc gagagggctc acccagcact gctgttagat acccctcagg cagtgtggcc 1200 ctcccaagcc agtggctgct cattggacac cacgagctac ccccagtcct gcacaccacc 1260 atgctgaggg tccatcccac cctggggagt ggaactgcag agacaagacc tccagagaat 1320 acccaggccc cagccttctt cttggagcta ttgagcctga gccgagagaa actttgggac 1380 tccgagctgc atccagaaga aaaaactcca gactcttact tggggctggg accccaagac 1440 ctgctggcag ctagcctcac tgctgtcctc ctgggagggt ggattctctt tgtgatgagg 1500 cagcagcagg agacccccct ggcacctgca gactttgctc acatctccca ggatgcccag 1560 tccctgcact cgggggccag ccggaggagc cagaagaggc ttcagagtcc ctcacctgag 1620 tcaccaccct cctctccccc agctgagcaa ctcaccgtag tggggaagat ttccttcaat 1680 cccaaggacg tgctgggccg cggggcaggc gggactttcg ttttcagggg acagtttgag 1740 ggacgggcag tggctgtcaa gcggctcctc cgcgagtgct ttggcctggt tcggcgggaa 1800 gttcaactgc tgcaggagtc tgacaggcac cccaacgtgc tccgctactt ctgcaccgag 1860 cggggacccc agttccacta cattgccctg gagctctgcc gggcctcctt gcaggagtac 1920 gtagaaaacc cggacctgga tcgcgggggt ctggagcccg aggtcgtgct gcagcagctg 1980 atgtctggcc tggcccacct gcactcttta cacatagtgc accgggacct gaagccagga 2040 aatattctca tcaccgggcc tgacagccag ggcctgggca gagtggtgct ctcagacttc 2100 ggcctctgca agaagctgcc tgctggccgc tgtagcttca gcctccactc cggcatcccc 2160 ggcacggaag gctggatggc gcccgagctt ctgcagctcc tgccaccaga cagtcctacc 2220 agcgctgtgg acatcttctc tgcaggctgc gtgttctact acgtgctttc tggtggcagc 2280 cacccctttg gagacagtct ttatcgccag gcaaacatcc tcacaggggc tccctgtctg 2340 gctcacctgg aggaagaggt ccacgacaag gtggttgccc gggacctggt tggagccatg 2400 ttgagcccac tgccgcagcc acgcccctct gccccccagg tgctggccca ccccttcttt 2460 tggagcagag ccaagcaact ccagttcttc caggacgtca gtgactggct ggagaaggag 2520 tccgagcagg agcccctggt gagggcactg gaggcgggag gctgcgcagt ggtccgggac 2580 aactggcacg agcacatctc catgccgctg cagacagatc tgagaaagtt ccggtcctat 2640 aaggggacat cagtgcgaga cctgctccgt gctgtgagga acaagaagca ccactacagg 2700 gagctcccag ttgaggtgcg acaggcactc ggccaagtcc ctgatggctt cgtccagtac 2760 ttcacaaacc gcttcccacg gctgctcctc cacacgcacc gagccatgag gagctgcgcc 2820 tctgagagcc tcttcctgcc ctactacccg ccagactcag aggccaggag gccatgccct 2880 ggggccacag ggaggtgagg tgggctggat gccacacaga tggtctccgt gctggctcac 2940 tgaagagctg agcctgtggc tggcctcaga atcaggctgg gtgcagtggc tcacacctgt 3000 aatcccagca ttttgggagg ctgagtgaga ggatcacttg agctcaggag ttcgagacca 3060 r gcctggccaa catggcaaca ccccatttct acaaaaaatt tgtaaaatta gccaggcatg 3120 gtggcgcacg cctgtagtcc cagctgcttg ggaggctgag gtgggagaat cacttgagcc 3180 caggagttcg aggctgcagt gagccaggat catgccactg cactccagcc tggtccacag 3240 agagacactg tcaccccctt tcccccacaa gactggcaga ggctgggcag cctggggctg 3300 atgaagcaga gatgttcgct ggatcccagc tcctggcaca ctgtaaggaa atacaacgaa 3360 gaggt 3365 <210> 34 <211> 2049 <212> DNA

<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 1851973CB1 <400> 34 gcgttctttc ccgcggaagt agttgacatt tacaaggagc agcgccccca aaggtcttta 60 gctgtttttt aaggggagaa cagcctttac cctctttgga ctttttcttc gttttttttt 120 tttttggaga cggagtttcg ttctttcgcc caggctggcg tacagtggcg cgatctcggc 180 tcactgcaac ctctgctccc cgggttcaag cgattctcct gcctcagcct ccggagtagc 240 tgggattaca ggtgcccgcc accacgcccg gctgatttcc tcttaagact ttctacagct 300 tccttatgaa atcttctgac tgggccttga gcaataaggt cttttgctac aatttagtgc 360 tcttttcctc acactaaatc gaaaactctc cctgttggtc ctgatctgtt tcagtcaggc 420 aaattacatc ctgggaaaac gtcagatgac aggggaggcc actcgcttcc tgctcatcca 480 gtttcgacac tttctgtgct ttcattagct tccagacctc agccctggcc ctcgctttac 540 tgtacagtca gaactggttt ctacgcctcg cgagggtggg aggtcgtgta tgggaggagg 600 accgcttccc accagcctcg ttgggaagcc aggagaaatc tcttcaaatc ctgcgattca 660 gagtcaagtc ccagtcgtcc tttttctggt cggcccagaa ctgtttgtgc ctcctccctc 720 atgaggaatg atgtcagtgg ggccgcggtc gccgcccacg aagagtgtaa ggctgcgaag 780 tcggggcttt cccgacgccc cctccgtccg cgtctgcgta ggggaggtga cgagggcggg 840 gcgcggcggc ggggtgacgt cacggccgcg cgcggcgtgg gcggagcctc actttgaacc 900 cagttggcgg gaatggctgc tcgcggaggg gcagtgtacg c.ggggccgct gtaggctgtc 960 cagcgatgga tcccaccgcg ggaagcaaga aggagcctgg aggaggcgcg gcgactgagg 1020 agggcgtgaa taggatcgca gtgccaaaac cgccctccat tgaggaattc agcatagtga 1080 agcccattag ccggggcgcc ttcgggaaag tgtatctggg gcagaaaggc ggcaaattgt 1140 atgcagtaaa ggttgttaaa aaagcagaca tgatcaacaa aaatatgact catcaggtcc 1200 aagctgagag agatgcactg gcactaagca aaagcccatt cattgtccat ttgtattatt 1260 cactgcagtc tgcaaacaat gtctacttgg taatggaata tcttattggg ggagatgtca 1320 agtctctcct acatatatat ggttattttg atgaagagat ggctgtgaaa tatatttctg 1380 aagtagcact ggctctagac taccttcaca gacatggaat catccacagg gacttgaaac 1440 cggacaatat gcttatttct aatgagggtc atattaaact gacggatttt ggcctttcaa 1500 aagttacttt gaatagagat attaatatga tggatatcct tacaacacca tcaatggcaa 1560 aacctagaca agattattca agaaccccag gacaagtgtt atcgcttatc agctcgttgg 1620 gatttaacac accaattgca gaaaaaaatc aagaccctgc aaacatcctt tcagcctgtc 1680 tgtctgaaac atcacagctt tctcaaggac tcgtatgccc tatgtctgta gatcaaaagg 1740 acactacgcc ttattctagc aaattactaa aatcatgtct tgaaacagtt gcctccaacc 1800 caggaatgcc tgtgaagtgt ctaacttcta atttactcca gtctaggaaa aggctggcca 1860 catccagtgc cagtagtcaa tcccacacct tcatatccag tgtggaatca gaatgccaca 1920 gcagtcccaa atgggaaaaa gattgccagg tttgagggac atttatctta atgaaaatca 1980 attatgtatg tcaaatgaat gtgagaaata ttataccttt tcatataaat tccataaaga 2040 aatgaaagg 2049 <210> 35 <211> 2962 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 7474604CB1 <400> 35 accactcgtg cccactgatt atcagcatct tttactttca ccagcgtttc tgggtgtcca 60 cctcctgcgg ccgcggcgga aaacatgacg aaaagcgagg agcagcagcc tctgagtttg 120 caaaaagcct tacagcagtg cgaactggtc caaaacatga tagacttgag catctccaac 180 ctggaagggc ttaggaccaa atgtgctacc tccaacgacc tcacacaaaa agaaatccgg 240 accctggaga gcaagctggt gaagtacttc agccggcagc tgtcctgcaa aaagaaggta 300 gccttgcagg agcgcaacgc ggagctggac ggcttccccc agctacggca ctggttccga 360 atcgtcgatg tgcgcaagga ggtcctggag gaaatctccc ccggccagct gagcctggag 420 gacctcttgg agatgacgga tgaacaggtg tgcgagactg tggagaaata cggagccaac 480 cgggaggagt gtgcccgcct caacgcctcc ctctcctgcc tcaggaatgt ccacatgtca 540 ggaggcaacc tttccaaaca agactggacc atccagtggc ccaccacaga gacggggaag 600 gagaacaatc ccgtgtgccc cccggagccc accccgtgga tccgcaccca tctctcccag 660 agccccaggg tcccgtccaa gtgcgtccag cactattgtc acaccagccc cactcccggg 720 gcccctgtgt acacccacgt ggacaggctt accgtggacg cctacccggg cttgtgcccg 780 cccccgccac tggagtcggg ccaccgttcc ctgcccccat cgccccggca gcggcacgcg 840 gtccgcaccc cgccgcgcac ccccaacatc gtcaccaccg tgaccccgcc gggcacgccg 900 cccatgagga agaagaacaa gctgaagccc ccggggaccc caccgccctc ctcccgaaaa 960 ctgatacact tgatcccggg attcaccgcg ctgcatcgga gcaaatccca cgagttccag 1020 ctggggcacc gcgtggacga ggcccacacg cccaaagcca agaagaagag caaacccttg 1080 aacctcaaga tccacagcag cgtaggcagc tgcgagaaca tcccctctca gcagcgctcc 1140 ccgctgctgt ccgagcgctc cctccgctcc ttctttgtgg gacacgcacc tttcctgcct 1200 tccacccctc ctgttcacac tgaggccaac ttctctgcaa acacactgtc agtgccacgc 1260 tggtccccgc agatccctcg cagagatctt ggcaactcca tcaagcacag gttttccacc 1320 aagtactgga tgtctcagac gtgcacagtc tgtgggaaag ggatgctttt tggcctcaag 1380 tgtaaaaact gcaagttaaa gtgccacaac aaatgcacca aagaagcccc accctgtcat 1440 cttctgatca tccaccgagg agatccagca aggttagtcc ggacagagtc cgttccgtgt 1500 gacatcaaca accctctacg gaagccacct cgctattcag acctgcacat cagtcagacg 1560 ctccccaaaa ccaacaaaat caacaaggac cacatccctg tcccttacca gccagactcc 1620 agcagcaacc cctcctccac gacgtcctcc acgccctcct cgccagcacc ccccctccct 1680 cctagtgcca cgccgccttc tcccctacac ccttccccac agtgcacacg gcagcagaag 1740 aacttcaacc tgccagcatc ccactactac aaatacaagc agcagttcat cttcccagat 1800 gtggtgccgg tgccggagac gccgacccgg gcgccccagg tcatcctgca tccggtgacc 1.860 tcgaatccaa tcttggaagg aaatccatta cttcaaattg aagtggagcc aacgtcggag 1920 aatgaagagg tccatgatga ggccgaagag tcagaggatg acttcgagga gatgaacctg 1980 tccctcctct cggcccggag cttcccacgc aaggccagcc agaccagcat cttccttcag 2040 gagtgggaca tcccctttga gcagctggag atcggcgagc tcattggaaa gggccgcttt 2100 gggcaagtgt accacggccg ctggcatggc gaggtggcca tccggctgat tgacattgag 2160 agggacaacg aggaccagct caaggccttc aagcgggagg tgatggccta caggcagaca 2220 cggcatgaga acgtggtgct tttcatgggt gcctgcatga gcccgcctca cctggccatc 2280 atcaccagcc tctgtaaggg acggacgctc tattccgttg tgagggatgc caaaatcgtt 2340 ttggatgtca acaaaaccag gcagattgct caagaaattg tgaagggcat gggctacctc 2400 cacgccaagg gaatcctaca caaggacctc aagtcaaaga acgtcttcta tgacaacggc 2460 aaagtggtca tcacggactt tggactcttc agcatttctg gggtgctgca ggctggcagg 2520 cgggaggaca aactgcgcat ccagaatggc tggctatgcc acctggcacc agagatcatc 2580 cgccagctgt cccccgacac agaggaggat aagctcccct tctccaagca ctctgacgtc 2640 tttgcccttg gcacaatctg gtatgaactc cacgccaggg aatggccttt caagacccaa 2700 ccagcagagg caataatctg gcaaatgggc acaggcatga aacccaacct cagccagatt 2760 ggcatgggaa aagaaatctc ggacattctt ctcttctgct gggcctttga acaagaagag 2820 agacctacct tcaccaagct catggacatg ctggagaaac tgccaaagcg aaaccgtcgc 2880 ctgtctcacc ctggacattt ctggaagtct gcagagctgt gacctttgga catcgggacg 2940 gcgcccagct gcctgggctc cc 2962 <210> 36 <211> 3112 <212> DNA
<213> Homo sapiens <220>
<221> misc_feature <223> Incyte ID No: 7474721CB1 <400> 36 gggggcattg ctcagcggtg ctaggctggc gcggcttgag ccgccgccgg actgacagct 60 cggtctgcgg accatggaga cctgcgccgg tccacacccg ctgcgcctct tcctctgccg 120 gatgcagctc tgtctcgcgc tgcttttggg accctggcgg cctgggaccg ccgaggaagt 180 tatcctcctg gattccaaag cctcccaggc cgagctgggc tggactgcac tgccaagtaa 240 tgggtgggag gagatcagcg gcgtggatga acacgaccgt cccatccgca cgtaccaagt 300 gtgcaatgtg ctggagccca accaggacaa ctggctgcag actggctgga taagccgtgg 360 ccgcgggcag cgcatcttcg tggaactgca gttcacactc cgtgactgca gcagcatccc 420 tggcgccgcg ggtacctgca aggagacctt caacgtctac tacctggaaa ctgaggccga 480 cctgggccgt gggcgtcccc gcctaggcgg cagccggccc cgcaaaatcg acacgatcgc 540 ggcggacgag agcttcacgc agggcgacct gggtgagcgc aagatgaagc tgaacacaga 600 ggtgcgcgag atcggaccgc tcagccggcg gggtttccac ctggcctttc aggacgtggg 660 cgcatgcgtg gcgcttgtct cggtgcgcgt ctactacaag cagtgccgcg ccaccgtgcg 720 gggcctggcc acgttcccag ccaccgcagc cgagagcgcc ttctccacac tggtggaagt 780 ggccggaacg tgcgtggcgc actcggaagg ggagcctggc agccccccac gcatgcactg 840 cggcgccgac ggcgagtggc tggtgcctgt gggccgctgc agctgcagcg cgggattcca 900 ggagcgtggt gacatctgcg aagcctgtcc cccagggttt tacaaggtgt ccccgcggcg 960 aagggtctgc tcaccgtgcc cagagcacag ccgggccctg gaaaacgcct ccaccttctg 1020 cgtgtgccag gacagctatg cgcgctcacc caccgacccg ccctcggctt cctgcacccg 1080 tgggccgccg tcggcgccgc gggacctgca gtacagcctg agccgctcgc cgctggtgct 1140 gcgactgcgc tggctgccgc cggccgactc gggaggccgc tcggacgtca cctactcgct 1200 gctgtgcctg cgctgcggcc gcgagggccc ggcgggcgcc tgcgagccgt gcgggccgcg 1260 cgtggccttc ctaccgcgcc aggcagggct gcgggagcga gccgccacgc tgctgcacct 1320 gcggcccggg gcgcgctaca ccgtgcgcgt ggccgtgctc aacggcgtct cgggcccggc 1380 ggccgccctg gttccggttg gcgctgtttc aattaaccct ggtacggttg gccctgttcc 1440 tgttgccggg gttatccgcg accgagtgga accccagagc gtgtccctgt cgtggcggga 1500 gcccatccct gccggagccc ctggggccaa tgacacggag tacgagatcc gatactacga 1560 gaaggtgcag agtgagcaga cttactccat ggtgaagaca ggggcgccca cagtcaccgt 1620 caccaacctg aagccggcta cccgctacgt ctttcagatc cgggccgctt ccccggggcc 1680 atcctgggag gcccagagtt ttaaccccag cattgaagta cagaccctgg gggaggctgc 1740 ctcagggtcc agggaccaga gccccgccat tgtcgtcacc gtagtgacca tctcggccct 1800 cctcgtcctg ggctccgtga tgagtgtgct ggccatttgg aggaggccct gcagctatgg 1860 caaaggagga ggggatgccc atgatgaaga ggagctgtat ttccacttca aagtcccaac 1920 acgtcgcaca ttcctggacc cccagagctg tggggacctg ctgcaggctg tgcatctgtt 1980 cgccaaggaa ctggatgcga aaagcgtcac gctggagagg agccttggag gagggcggtt 2040 tggggagctg tgctgtggct gcttgcagct ccccggtcgc caggagctgc tcgtagccgt 2100 gcacatgctg agggacagcg cctccgactc acagaggctc ggcttcctgg ccgaggccct 2160 cacgctgggc cagtttgacc atagccacat cgtgcggctg gagggcgttg ttacccgagg 2220 aagcaccttg atgattgtca ccgagtacat gagccatggg gccctggacg gcttcctcag 2280 gcggcacgag gggcagctgg tggctgggca actgatgggg ttgctgcctg ggctggcatc 2340 agccatgaag tatctgtcag agatgggcta cgttcaccgg ggcctggcag ctcgccatgt 2400 gctggtcagc agcgaccttg tctgcaagat ctctggcttc gggcggggcc cccgggaccg 2460 atcagaggct gtctacacca ctatgagtgg ccggagccca gcgctatggg ccgctcccga 2520 gacacttcag tttggccact tcagctctgc cagtgacgtg tggagcttcg gcatcatcat 2580 gtgggaggtg atggcctttg gggagcggcc ttactgggac atgtctggcc aagacgtgat 2640 caaggctgtg gaggatggct tccggctgcc accccccagg aactgtccta accttctgca 2700 ccgactaatg ctcgactgct ggcagaagga cccaggtgag cggcccaggt tctcccagat 2760 ccacagcatc ctgagcaaga tggtgcagga cccagagccc cccaagtgtg ccctgactac 2820 ctgtcccagg cctcccaccc cactagcgga ccgtgccttc tccaccttcc cctcctttgg 2880 ctctgtgggc gcgtggctgg aggccctgga cctgtgccgc tacaaggaca gcttcgcggc 2940 tgctggctat gggagcctgg aggccgtggc cgagatgact gcccagaggg acctggtgag 3000 cctaggcatc tctttggctg aacatcgaga ggccctcctc agcgggatca gcgccctgca 3060 ggcacgagtg ctccagctgc agggccaggg ggtgcaggtg tgagtggacc cc 3112 <210> 37 <211> 3650 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 7478815CB1 <400> 37 caacactcca gagtcgtagg agtgaacact gcacaggaat ctctgcccat ctcaggagaa 60 accaaacttg gggaaaatgt ttgcggtcca cttgatggca ttttacttca gcaagctgaa 120 ggaggaccag atcaagaagg tggacaggtt cctgtatcac atgcggctct ccgatgacac 180 ccttttggac atcatgaggc ggttccgggc tgagatggag aagggcctgg caaaggacac 240 caaccccacg gctgcagtga agatgttgcc caccttcgtc agggccattc ccgatggttc 300 cgaaaatggg gagttccttt ccctggatct cggagggtcc aagttccgag tgctgaaggt 360 gcaagtcgct gaagagggga agcgacacgt gcagatggag agtcagttct acccaacgcc 420 caatgaaatc atccgcggga acggcacaga gctgtttgaa tatgtagctg actgtctggc 480 agatttcatg aagaccaaag atttaaagca taagaaattg ccccttggcc taactttttc 540 tttcccctgt cgacagacta aactggaaga gggtgtccta ctttcgtgga caaaaaagtt 600 taaggcacga ggagttcagg acacggatgt ggtgagccgt ctgaccaaag ccatgagaag 660 acacaaggac atggacgtgg acatcctggc cctggtcaat gacaccgtgg ggaccatgat 720 gacctgtgcc tatgacgacc cctactgcga agttggtgtc atcatcggaa ctggcaccaa 780 tgcgtgttac atggaggaca tgagcaacat tgacctggtg gagggcgacg agggcaggat 840 gtgcatcaac acagagtggg gggccttcgg ggacgacggg gccctggagg acattcgcac 900 tgagttcgac agggagctgg acctcggctc tctcaaccca ggaaagcaac tgttcgagaa 960 gatgatcagt ggcctgtacc tgggggagct tgtcaggctt atcttgctga agatggccaa 1020 ggctggcctc ctgtttggtg gtgagaaatc ttctgctctc cacactaagg gcaagatcga 1080 aacacggcac gtggctgcca tggagaagta taaagaaggc cttgctaata caagagagat 1140 cctggtggac ctgggtctgg aaccgtctga ggctgactgc attgccgtcc agcatgtctg 1200 taccatcgtc tccttccgct cggccaatct ctgtgcagca gctctggcgg ccatcctgac 1260 acgcctccgg gagaacaaga aggtggaacg gctccggacc acagtgggca tggacggcac 1320 cctctacaag atacaccctc agtacccaaa acgcctgcac aaggtggtga ggaaactggt 1380 cccaagctgt gatgtccgct tcctcctgtc agagagtggc agcaccaagg gggccgccat 1440 ggtgaccgcg gtggcctccc gcgtgcaggc ccagcggaag cagatcgaca gggtgctggc 1500 tttgttccag ctgacccgag agcagctcgt ggacgtgcag gccaagatgc gggctgagct 1560 ggagtatggg ctgaagaaga agagccacgg gctggccacg gtcaggatgc tgcccaccta 1620 cgtctgcggg ctgccggacg gcacagagaa aggaaagttt ctcgccctgg atcttggggg 1680 aaccaacttc cgggtcctcc tggtgaagat cagaagtgga cggaggtcag tgcgaatgta 1740 caacaagatc ttcgccatcc ccctggagat catgcagggc actggtgagg agctctttga 1800 tcacattgtg cagtgcatcg ccgacttcct ggactacatg ggcctcaagg gagcctccct 1860 acctttgggc ttcacattct catttccctg caggcagatg agcattgaca agggaacact 1920 catagggtgg accaaaggtt tcaaggccac tgactgtgaa ggggaggacg tggtggacat 1980 gctcagggaa gccatcaaga ggagaaacga gtttgacctg gacattgttg cagtcgtgaa 2040 tgatacagtg gggaccatga tgacctgtgg ctatgaagat cctaattgtg agattggcct 2100 gattgcagga acaggcagca acatgtgcta catggaggac atgaggaaca tcgagatggt 2160 ggaggggggt gaagggaaga tgtgcatcaa tacagagtgg ggaggatttg gagacaatgg 2220 ctgcatagat gacatccgga cccgatacga cacggaggtg gatgaggggt ccttgaatcc 2280 tggcaagcag agatacgaga aaatgaccag tgggatgtac ttgggggaga ttgtgcggca 2340 gatcctgatc gacctgacca agcagggtct cctcttccga gggcagattt cagagcgtct 2400 ccggaccagg ggcatcttcg aaaccaagtt cctgtcccag atcgaaagcg atcggctggc 2460 ccttctccag gtcaggagga ttctgcagca gctgggcctg gacagcacgt gtgaggacag 2520 catcgtggtg aaggaggtgt gcggagccgt gtcccggcgg gcggcccagc tctgcggtgc 2580 tggcctggcc gctatagtgg aaaaaaggag agaagaccag gggctagagc acctgaggat 2640 cactgtgggt gtggacggca ccctgtacaa gctgcaccct cacttttcta gaatattgca 2700 ggaaactgtg aaggaactag cccctcgatg tgatgtgaca ttcatgctgt cagaagatgg 2760 cagtggaaaa ggggcagcac tgatcactgc tgtggccaag aggttacagc aggcacagaa 2820 ggagaactag gaacccctgg gattggacct gatgcatctt ggatactgaa cagcttttcc 2880 tctggcagat cagttggtca gagaccaatg ggcaccctcc tggctgacct caccttctgg 2940 atggccgaaa gagaacccca ggttctcggg tactcttagt atcttgtact ggatttgcag 3000 tgacattaca tgacatctct atttggtata tttgggccaa aatgggccaa cttatgaaat 3060 caaagtgtct gtcctgagag atcccctttc aacacattgt tcaggtgagg cttgagctgt 3120 caattctcta tggctttcag tcttgtggct gcgggacttg gaaatatata gaatctgccc 3180 atgtggctgg caggctgttt ccccattggg atgcttaagc catctcttat aggggattgg 3240 accctgtact tgtggatgaa cattggagag caagaggaac tcacgttatg aactaggggg 3300 atctcatcta acttgtcctt aacttgccat gttgacttca aacctgttaa gagaacaaag 3360 actttgaagt atccagcccc agggtgcaga gaggttgatt gccagggagc actgcaggaa 3420 tcattgcatg cttaaagcga gttatgtcag caccctgtag gattttgttc cttattaagt 3480 gtgtgccatg tggtggggtg ctgtctgggg catctgtttt tcattttgcc tgtggtttgt 3540 gttgcaggtg ttgatagttg ttttaaggat tgttaggtat aggaaatcca gtaaattaat 3600 aaaaaaattt tgattttcca ataaaaaaaa aaaaaaaaaa aaaaaaaaaa 3650 <210> 38 <211> 7789 <212> DNA
<213> Homo Sapiens <220>
<221> misc feature <223> Incyte ID No: 7477141CB1 <400> 38 cacaccctga aagccggtcc ctggccgtgc tggcccccct gcaggacgtg gacgtggggg 60 ccggggagat ggcgctgttt gagtgcctgg tggcggggcc cactgacgtg gaggtggatt 120 ggctgtgccg tggccgcctg ctgcagcctg cactgctcaa atgcaagatg catttcgatg 180 gccgcaaatg caagctgcta cttacatctg tacatgagga cgacagtggc gtctacacct 240 gcaagctcag cacggccaaa gatgagctga cctgcagtgc ccggctgacc gtgcggccct 300 cgttggcacc cctgttcaca cggctgctgg aagatgtgga ggtgttggag ggccgagctg 360 cccgtttcga ctgcaagatc agtggcaccc cgccccctgt tgttacctgg actcattttg 420 gctgccccat ggaggagagt gagaacttgc ggctgcggca ggacgggggt ctgcactcac 480 tgcacattgc ccatgtgggc agcgaggacg aggggctcta tgcggtcagt gctgttaaca 540 cccatggcca ggcccactgc tcagcccagc tgtatgtaga agagccccgg acagccgcct 600 caggccccag ctcgaagctg gagaagatgc catccattcc cgaggagcca gagcagggtg 660 agctggagcg gctgtccatt cccgacttcc tgcggccact gcaggacctg gaggtgggac 720 tggccaagga ggccatgcta gagtgccagg tgaccggcct gccctacccc accatcagct 780 ggttccacaa tggccaccgc atccagagca gcgacgaccg gcgcatgaca cagtacaggg 840 atgtccatcg cttggtgttc cctgccgtgg ggcctcagca cgccggtgtc tacaagagcg 900 tcattgccaa caagctgggc aaagctgcct gctatgccca cctgtatgtc acagatgtgg 960 tcccaggccc tccagatggc gccccgcagg tggtggctgt gacggggagg atggtcacac 1020 tcacatggaa cccccccagg agtctggaca tggccatcga cccggactcc ctgacgtaca 1080 cagtgcagca ccaggtgctg ggctcggacc agtggacggc actggtcaca ggcctgcggg 1140 agccagggtg ggcagccaca gggctgcgta agggggtcca gcacatcttc cgggtcctca 1200 gcaccactgt caagagcagc agcaagccct cacccccttc tgagcctgtg cagctgctgg 1260 agcacggccc aaccctggag gaggcccctg ccatgctgga caaaccagac atcgtgtatg 1320 tggtggaggg acagcctgcc agcgtcaccg tcacattcaa ccatgtggag gcccaggtcg 1380 tctggaggag ctgccgaggg gccctcctag aggcacgggc cggtgtgtac gagctgagcc 1440 agccagatga tgaccagtac tgtcttcgga tctgccgggt gagccgccgg gacatggggg 1500 ccctcacctg caccgcccga aaccgtcacg gcacacagac ctgctcggtc acattggagc 1560 tggcagaggc ccctcggttt gagtccatca tggaggacgt ggaggtgggg gctggggaaa 1620 ctgctcgctt tgcggtggtg gtcgagggaa aaccactgcc ggacatcatg tggtacaagg 1680 acgaggtgct gctgaccgag agcagccatg tgagcttcgt gtacgaggag aatgagtgct 1740 ccctggtggt gctcagcacg ggggcccagg atggaggcgt ctacacctgc accgcccaga 1800 acctggcggg tgaggtctcc tgcaaagcag agttggctgt gcattcagct cagacagcta 1860 tggaggtcga gggggtcggg gaggatgagg accatcgagg aaggagactc agcgactttt 1920 atgacatcca ccaggagatc ggcaggggtg ctttctccta cttgcggcgc atagtggagc 1980 gtagctccgg cctggagttt gcggccaagt tcatccccag ccaggccaag ccaaaggcat 2040 cagcgcgtcg ggaggcccgg ctgctggcca ggctccagca cgactgtgtc ctctacttcc 2100 atgaggcctt cgagaggcgc cggggactgg tcattgtcac cgagctctgc acagaggagc 2160 tgctggagcg aatcgccagg aaacccaccg tgtgtgagtc tgagatccgg gcctatatgc 2220 ggcaggtgct agagggaata cactacctgc accagagcca cgtgctgcac ctcgatgtca 2280 agcctgagaa cctgctggtg tgggatggtg ctgcgggcga gcagcaggtg cggatctgtg 2340 actttgggaa tgcccaggag ctgactccag gagagcccca gtactgccag tatggcacac 2400 ctgagtttgt agcacccgag attgtcaatc agagccccgt gtctggagtc actgacatct 2460 ggcctgtggg tgttgttgcc ttcctctgtc tgacaggaat ctccccgttt gttggggaaa 2520 atgaccggac aacattgatg aacatccgaa actacaacgt ggccttcgag gagaccacat 2580 tcctgagcct gagcagggag gcccggggct tcctcatcaa agtgttggtg caggaccggc 2640 tgagacctac cgcagaagag accctagaac atccttggtt caaaactcag gcaaagggcg 2700 cagaggtgag cacggatcac ctgaagctat tcctctcccg gcggaggtgg cagcgctccc 2760 agatcagcta caaatgccac ctggtgctgc gccccatccc cgagctgctg cgggcccccc 2820 cagagcgggt gtgggtgacc atgcccagaa ggccaccccc cagtgggggg ctctcatcct 2880 cctcggattc tgaagaggaa gagctggaag agctgccctc agtgccccgc ccactgcagc 2940 ccgagttctc tggctcccgg gtgtccctca cagacattcc cactgaggat gaggccctgg 3000 ggaccccaga gactggggct gccaccccca tggactggca ggagcaggga agggctccct 3060 ctcaggacca ggaggctccc agcccagagg ccctcccctc cccaggccag gagcccgcag 3120 ctggggctag ccccaggcgg ggagagctcc gcaggggcag ctcggctgag agcgccctgc 3180 cccgggccgg gccgcgggag ctgggccggg gcctgcacaa ggcggcgtct gtggagctgc 3240 cgcagcgccg gagccccggc ccgggagcca cccgcctggc ccggggaggc ctgggtgagg 3300 gcgagtatgc ccagaggctg caggccctgc gccagcggct gctgcgggga ggccccgagg 3360 atggcaaggt cagcggcctc aggggtcccc tgctggagag cctggggggc cgtgctcggg 3420 acccccggat ggcacgagct gcctccagcg aggcagcgcc ccaccaccag cccccactcg 3480 agaaccgggg cctgcaaaag agcagcagct tctcccaggg tgaggcggag ccccggggcc 3540 ggcaccgccg agcgggggcg cccctcgaga tccccgtggc caggcttggg gcccgtaggc 3600 tacaggagtc tccttccctg tctgccctca gcgaggccca gccatccagc cctgcacggc 3660 ccagcgcccc caaacccagt acccctaagt ctgcagaacc ttctgccacc acacctagtg 3720 atgctccgca gccccccgca ccccagcctg cccaagacaa ggctccagag cccaggccag 37$0 aaccagtccg agcctccaag cctgcaccac ccccccaggc cctgcaaacc ctagcgctgc 3840 ccctcacacc ctatgctcag atcattcagt ccctccagct gtcaggccac gcccagggcc 3900 cctcgcaggg ccctgccgcg ccgccttcag agcccaagcc ccacgctgct gtctttgcca 3960 gggtggcctc cccacctccg ggagcccccg agaagcgcgt gccctcagcc gggggtcccc 4020 cggtgctagc cgagaaagcc cgagttccca cggtgccccc caggccaggc agcagtctca 4080 gtagcagcat cgaaaacttg gagtcggagg ccgtgttcga ggccaagttc aagcgcagcc 4140 gcgagtcgcc cctgtcgctg gggctgcggc tgctgagccg ttcgcgctcg gaggagcgcg 4200 gccccttccg tggggccgag gaggaggatg gcatataccg gcccagcccg gcggggaccc 4260 cgctggagct ggtgcgacgg cctgagcgct cacgctcggt gcaggacctc agggctgtcg 4320 gagagcctgg cctcgtccgc cgcctctcgc tgtcactgtc ccagcggctg cggcggaccc 4380 ctcccgcgca gcgccacccg gcctgggagg cccgcggcgg ggacggagag agctcggagg 4440 gcgggagctc ggcgcggggc tccccggtgc tggcgatgcg caggcggctg agcttcaccc 4500 tggagcggct gtccagccga ttgcagcgca gtggcagcag cgaggactcg gggggcgcgt 4560 cgggccgcag cacgccgctg ttcggacggc ttcgcagggc cacgtccgag ggcgagagtc 4620 tgcggcgcct tggccttccg cacaaccagt tggccgccca ggccggcgcc accacgcctt 4680 ccgccgagtc cctgggctcc gaggccagcg ccacgtcggg ctcctcagcc ccaggggaaa 4740 gccgaagccg gctccgctgg ggcttctctc ggccgcggaa ggacaagggg ttatcgccac 4800 caaacctctc tgccagcgtc caggaggagt tgggtcacca gtacgtgcgc agtgagtcag 4860 acttcccccc agtcttccac atcaaactca aggaccaggt gctgctggag ggggaggcag 4920 ccaccctgct ctgcctgcca gcggcctgcc ctgcaccgca catctcctgg atgaaagaca 4980 agaagtcctt gaggtcagag ccctcagtga tcatcgtgtc ctgcaaagat gggcggcagc 5040 tgctcagcat cccccgggcg ggcaagcggc acgccggtct ctatgagtgc tcggccacca 5100 acgtactggg cagcatcacc agctcctgta ccgtggctgt ggcccgagtc ccaggaaagc 5160 tagctcctcc agaggtaccc cagacctacc aggacacggc gctggtgctg tggaagccgg 5220 gagacagccg ggcaccttgc acgtatacgc tggagcggcg agtggatggg gagtctgtgt 5280 ggcaccctgt gagctcaggc atccccgact gttactacaa cgtgacccac ctgccagttg 5340 gcgtgactgt gaggttccgt gtggcctgtg ccaaccgtgc tgggcagggg cccttcagca 5400 actcttctga gaaggtcttt gtcaggggta ctcaagattc ttcagctgtg ccatctgctg 5460 cccaccaaga ggcccctgtc acctcaaggc cagccagggc ccggcctcct gactctccta 5520 cctcactggc cccaccccta gctcctgctg cccccacacc cccgtcagtc actgtcagcc 5580 cctcatctcc ccccacacct cctagccagg ccttgtcctc gctcaaggct gtgggtccac 5640 caccccaaac ccctccacga agacacaggg gcctgcaggc tgcccggcca gcggagccca 5700 ccctacccag tacccacgtc accccaagtg agcccaagcc tttcgtcctt gacactggga 5760 ccccgatccc agcctccact cctcaagggg ttaaaccagt gtcttcctct actcctgtgt 5820 atgtggtgac ttcctttgtg tctgcaccac cagcccctga gcccccagcc cctgagcccc 5880 ctcctgagcc taccaaggtg actgtgcaga gcctcagccc ggccaaggag gtggtcagct 5940 cccctgggag cagtccccga agctctccca ggcctgaggg taccactctt cgacagggtc 6000 cccctcagaa accctacacc ttcctggagg agaaagccag gggccgcttt ggtgttgtgc 6060 gagcgtgccg ggagaatgcc acggggcgaa cgttcgtggc caagatcgtg ccctatgctg 6120 ccgagggcaa gcggcgggtc ctgcaggagt acgaggtgct gcggaccctg caccacgagc 6180 ggatcatgtc cctgcacgag gcctacatca cccctcggta cctcgtgctc attgctgaga 6240 gctgtggcaa ccgggaactc ctctgtgggc tcagtgacag gttccggtat tctgaggatg 6300 acgtggccac ttacatggtg cagctgctac aaggcctgga ctacctccac ggccaccacg 6360 tgctccacct agacatcaag ccagacaacc tgctgctggc ccctgacaat gccctcaaga 6420 ttgtggactt tggcagtgcc cagccctaca acccccaggc ccttaggccc cttggccacc 6480 gcacgggcac gctggagttc atggctccgg agatggtgaa gggagaaccc atcggctctg 6540 ccacggacat ctggggagcg ggtgtgctca cttacattat gctcagtgga cgctccccgt 6600 tctatgagcc agacccccag gaaacggagg ctcggattgt ggggggccgc tttgatgcct 6660 tccagctgta ccccaataca tcccagagcg ccaccctctt cttgcgaaag gttctctctg 6720 tacatccctg gagccggccc tccctgcagg actgcctggc ccacccatgg ttgcaggacg 6780 cctacctgat gaagctgcgc cgccagacgc tcaccttcac caccaaccgg ctcaaggagt 6840 tcctgggcga gcagcggcgg cgccgggctg aggctgccac ccgccacaag gtgctgctgc 6900 gctcctaccc tggcggcccc tagaggcacg gaccacagcc aggcctcggg cttcaactgg 6960 ggttcccacc aatgccacgg gacattccag ggcccacgct gagccaggcg ggcctggggc 7020 ttcggttacc accagcagca acatctggct gggctcttac ctcatagacc ttcaaggaca 7080 gagaccccag ggcctggacc tgatgccacc ccaggccaaa gccagagtgg gagacccatt 7140 ggtcaggctc agcagggtgg gaacaggcag agggacaaga ggggaatgga gaagtggaga 7200 ggaaaaggaa tcgagggaca ggaaggggga ggctctagga aggttctggg ttgggggtca 7260 gtgcatctca gggagaacca aggaaggtgg gcatggctgg agaggaggaa aaggaaggag 7320 ccccaggtgt cagggcagta ggctgggagt cagtgtggca aagcgggggc aggacacaga 7380 tacagtggca ggggcccagg gctgggacat gagagaaggc agcgaggcgg cagagggaga 7440 agagaggact caggtggagg tggggtgggt cagctgtcag catccctcag aggagaaatg 7500 tggagagctg gaggccagca gtcactcaca ctcgctctgt cctcctgtcc agtggataca 7560 gccctgggcg ctctgctggc ccaaggatgt ccccactgcc cctccatggc ctttggcctt 7620 cttcccattc atatttattt atttattgac ttttatgaag tttccccttc catccgatcc 7680 ctactgccca tgttgtcctg accatccctc ccagccatcc agctgtctgt ctgtctgcca 7740 caaggaaata aaaatggcaa gcagcataaa aaaaaaaaaa aaaaaaaaa 7789 <210> 39 <211> 1937 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte TD No: 2190612CB1 <400> 39 gtggtgtggc tgcagtggag agttcccaac aaggctacgc agaagaaccc ccttgactga 60 agcaatggag gggggtccag ctgtctgctg ccaggatcct cgggcagagc tggtagaacg 120 ggtggcagcc atcgatgtga ctcacttgga ggaggcagat ggtggcccag agcctactag 180 aaacggtgtg gaccccccac cacgggccag agctgcctct gtgatccctg gcagtacttc 240 aagactgctc ccagcccggc ctagcctctc agccaggaag ctttccctac aggagcggcc 300 agcaggaagc tatctggagg cgcaggctgg gccttatgcc acggggcctg ccagccacat 360 ctccccccgg gcctggcgga ggcccaccat cgagtcccac cacgtggcca tctcagatgc 420 agaggactgc gtgcagctga accagtacaa gctgcagagt gagattggca agggtgccta 480 cggtgtggtg aggctggcct acaacgaaag tgaagacaga cactatgcaa tgaaagtcct 540 ttccaaaaag aagttactga agcagtatgg ctttccacgt cgccctcccc cgagagggtc 600 ccaggctgcc cagggaggac cagccaagca gctgctgccc ctggagcggg tgtaccagga 660 gattgccatc ctgaagaagc tggaccacgt gaatgtggtc aaactgatcg aggtcctgga 720 tgacccagct gaggacaacc tctatttggt gtttgacctc ctgagaaagg ggcccgtcat 780 ggaagtgccc tgtgacaagc ccttctcgga ggagcaagct cgcctctacc tgcgggacgt 840 catcctgggc ctcgagtact tgcactgcca gaagatcgtc cacagggaca tcaagccatc 900 caacctgctc ctgggggatg atgggcacgt gaagatcgcc gactttggcg tcagcaacca 960 gtttgagggg aacgacgctc agctgtccag cacggcggga accccagcat tcatggcccc 1020 cgaggccatt tctgattccg gccagagctt cagtgggaag gccttggatg tatgggccac 1080 tggcgtcacg ttgtactgct ttgtctatgg gaagtgcccg ttcatcgacg atttcatcct 1140 ggccctccac aggaagatca agaatgagcc cgtggtgttt cctgaggagc cagaaatcag 1200 cgaggagctc aaggacctga tcctgaagat gttagacaag aatcccgaga cgagaattgg 1260 ggtgccagac atcaagttgc acccttgggt gaccaagaac ggggaggagc cccttccttc 1320 ggaggaggag cactgcagcg tggtggaggt gacagaggag gaggttaaga actcagtcag 1380 gctcatcccc agctggacca cggtgatcct ggtgaagtcc atgctgagga agcgttcctt 1440 tgggaacccg tttgagcccc aagcacggag ggaagagcga tccatgtctg ctccaggaaa 1500 cctactggtg aaagaagggt ttggtgaagg gggcaagagc ccagagctcc ccggcgtcca 1560 ggaagacgag gctgcatcct gagcccctgc atgcacccag ggccacccgg cagcacactc 1620 atcccgcgcc tccagaggcc cacccctcat gcaacagccg cccccgcagg cagggggctg 1680 gggactgcag ccccactccc gcccctcccc catcgtgctg catgacctcc acgcacgcac 1740 gtccagggac agactggaat gtatgtcatt tggggtcttg ggggcagggc tcccacgagg 1800 ccatcctcct cttcttggac ctccttggcc tgagccattc tgtggggaaa ccgggtgccc 1860 atggagcctc agaaatgaca cccggctggt tggcatggcc tggggcagga ggcagaggca 1920 ggagaccaag atggcag 1937 <210> 40 <211> 5373 <212> DNA
<213> Homo Sapiens <220>
<221> misc_feature <223> Incyte ID No: 7477549CB1 <400> 40 atggagcggc ggctgcgcgc gctggagcag ctggcgcggg gcgaggccgg cggctgcccg 60 gggctcgacg gcctcctaga tctgctgctg gcgctgcacc acgagctcag cagcggcccc 120 ctacggcggg agcgcagcgt ggcgcagttc ctgagctggg ccagcccctt cgtatcaaag 180 gtgaaagaac tgcgtctgca gagagatgac tttgagatct tgaaggtgat cggccgagga 240 gcctttgggg aggtcaccgt ggtgaggcag agggacactg ggcagatttt tgccatgaaa 300 atgctgcaca agtgggagat gctgaagagg gctgagacag cctgtttcdg ggaggagcgg 360 gatgtgctcg tgaaagggga cagccgttgg gtgaccactc tgcactatgc cttccaagac 420 gaggagtacc tgtaccttgt gatggactac tatgctggtg gggacctcct gacgctgctg 480 agccgcttcg aggaccgtct cccgcccgag ctggeccagt tctacctggc tgagatggtg 540 ctggccatcc actcgctgca ccagctgggt tatgtccaca gggatgtcaa gccagacaac 600 gtcctgctgg atgtgaacgg gcacattcgc ctggctgact tcggctcctg cctgcgtctc 660 aacaccaacg gcatggtgga ttcatcagtg gcagtaggga cgccggacta tatctcccct 720 gagatcctgc aggccatgga ggagggcaag ggccactacg gcccacagtg tgactggtgg 780 tcgcttggag tctgcgccta tgagctgctc tttggggaga cgcccttcta tgctgagtcc 840 ttggtggaaa cctacggcaa gatcatgaac cacgaggacc acctgcagtt ccccccggac 900.

gtgcctgacg tgccagccag cgcccaagac ctgatccgcc agctgctgtg tcgccaggaa 960 gagcggctag gccgtggtgg gctggatgac ttccggaacc atcctttctt cgaaggcgtg 1020 gactgggagc ggctggcgag cagcacggcc ccctatattc ctgagctgcg ggggcccatg 1080 gacacctcca actttgatgt ggatgacgac accctcaacc atccagggac cctgccaccg 1140 ccctcccacg gggccttctc cggccatcac ctgccattcg tgggcttcac ctacacctca 1200 ggcagtcaca gtcctgagag cagctctgag gcttgggctg ccctggagcg gaagctccag 1260 tgtctggagc aggagaaggt ggagctgagc aggaagcacc aagaggccct gcacgccccc 1320 acagaccatc gggagctgga gcagctacgg aaggaagtgc agactctgcg ggacaggctg 1380 ccagagatgc tgagggacaa ggcctcattg tcccagacgg atgggccccc agctggtagc 1440 ccaggtcagg acagtgacct acggcaggag cttgaccgac ttcaccggga gctggccgag 1500 ggtcgggcag ggctgcaggc tcaggagcag gagctctgca gggcccaggg gcagcaggag 1560 gagctgcttc agaggctaca ggaggcccag gagagagagg cggccacagc tagccagacc 1620 cgggccctga gctcccagct ggaggaagcc cgggctgccc agagggagct ggaggcccag 1680 gtgtcctccc tgagccggca ggtgacgcag ctgcagggac agtgggagca acgccttgag 1740 gagtcgtccc aggccaagac catccacaca gcctctgaga ccaacgggat gggaccccct 1800 gagggtgggc ctcaggaggc ccaactgagg aaggaggtgg ccgccctgcg agagcagctg 1860 gagcaggccc acagccacag gccgagtggt aaggaggagg ctctgtgcca gctgcaggag 1920 gaaaaccgga ggctgagccg ggagcaggag cggctagaag cagagctggc ccaggagcag 1980 gagagcaagc agcggctgga gggtgagcgg cgggagacgg agagcaactg ggaggcccag 2040 ctcgccgaca tcctcagctg ggtgaatgat gagaaggtct caagaggcta cctgcaggcc 2100 ctggccacca agatggcaga ggagctggag tccttgagga acgtaggcac ccagacgctc 2160 cctgcccggc cactgaagat ggaggcctcg gccaggctgg agctgcagtc agcgctggag 2220 gccgagatcc gcgccaagca gggcctgcag gagcggctga cacaggtgca ggaggcccag 2280 ctgcaggctg agcgccgtct gcaggaggcc gagaagcaga gccaggccct gcaacaggag 2340 ctcgccatgc tgcgggagga gctgcgggcc cgagggccag tggacaccaa gccctcaaac 2400 tccctgattc ccttcctgtc cttccggagc tcagagaagg attctgccaa ggaccctggc 2460 atctcaggag aggccacaag gcatggagga gagccagatc tgaggccgga gggccgacgc 2520 agcctgcgca tgggggctgt gttccccaga gcacccactg ccaacacagc ctctacagaa 2580 ggtcttcctg ctaagggatg gggcatgggg ccctgggagg ccttgggtaa tggctgtccc 2640 cctccccagc ccggctcaca cacgctgcgc ccccggagct tcccatcccc gaccaagtgt 2700 ctccgctgca cctcgctgat gctgggcctg ggccgccagg gcctgggttg tgatgcctgc 2760 ggctactttt gtcacacaac ctgtgcccca caggccccac cctgccccgt gccccctgac 2820 ctcctccgca cagccctggg agtacacccc gaaacaggca caggcactgc ctatgagggc 2880 tttctgtcgg tgccgcggcc ctcaggtgtc cggcggggct ggcagcgcgt gtttgctgcc 2940 ctgagtgact cacgcctgct gctgtttgac gcccctgacc tgaggctcag cccgcccagt 3000 ggggccctcc tgcaggtcct agatctgagg gacccccagt tctcggctac ccctgtcctg 3060 gcctctgatg ttatccatgc ccaatccagg gacctgccac gcatctttag ggtgacaacc 3220 tcccagctgg cagtgccgcc caccacgtgc actgtgctgc tgctggcaga gagcgagggg 3180 gagcgggaac gctggctgca ggtgctgggt gagctgcagc ggctgctgct ggacgcgcgg 3240 ccaagacccc ggcccgtgta cacactcaag gaggcttacg acaacgggct gccgctgctg 3300 cctcacacgc tctgcgctgc catcctegac caggatcgac ttgcgcttgg caccgaggag 3360 gggctctttg tcatccatct gcgcagcaac gacatcttcc aggtggggga gtgccggcgc 3420 gtgcagcagc tgaccttgag ccccagtgca ggcctgctgg tcgtgctgtg tggccgcggc 3480 cccagcgtgc gtctctttgc cctggcggag ctggagaaca tagaggtagc aggtgccaag 3540 atccccgagt ctcgaggctg ccaggtgctg gcagctggaa gcatcctgca ggcccgcacc 3600 ccggtgctct gtgtagccgt caagcgccag gtgctctgct accagctggg cccgggccct 3660 gggccctggc agcgccgcat ccgtgagctg caggcacctg ccactgtgca gagcctgggg 3720 ctgctgggag accggctatg tgtgggcgcc gccggtggct ttgcactcta cccgctgctc 3780 aacgaggctg cgccgttggc gctgggggcc ggtttggtgc ctgaggagct gccaccatcc 3840 cgcgggggcc tgggtgaggc actgggtgcc gtggagctta gcctcagcga gttcctgcta 3900 ctcttcacca ctgctggcat ctacgtggat ggcgcaggcc gcaagtctcg tggccacgag 3960 ctgttgtggc cagcagcgcc catgggctgg gggtatgcgg ccccctacct gacagtgttc 4020 agcgagaact ccatcgatgt gtttgacgtg aggagggcag aatgggtgca gaccgtgccg 4080 ctcaagaagg tgcggcccct caatccagag ggctccctgt tcctctacgg caccgagaag 4140 gtccgcctga cctacctcag gaaccagctg gcagagaagg acgagttcga catcccggac 4200 ctcaccgaca acagccggcg ccagctgttc cgcaccaaga gcaagcgccg cttctttttc 4260 cgcgtgtcgg aggagcagca gaagcagcag cgcagggaga tgctgaagga cccttttgtg 4320 cgctccaagc tcatctcgcc gcctaccaac ttcaaccacc tagtacacgt gggccctgcc 4380 aacgggcggc ccggcgccag ggacaagtcc ccgtcccagc ccctccgcac tgtcacccaa 4440 caggctcccg aagagaaggg ccgagttgcc cgcggctccg gcccacagcg gccccacagc 4500 ttctccgagg cgttgcggcg cccagcctcc atgggcagcg aaggcctcgg tggagacgca 4560 gaccccactg gagcagtgaa gaggaaaccc tggacatccc tgtccagcga gtctgtgtcc 4620 tgcccccagg gatcgctgag ccctgcaacc tccctaatgc aggtctcaga acggccccga 4680 agcctccccc tgtcccctga attggagagc tctccttgat gccctctgtt agggcccacc 4740 ccaatcccag ggcagaagga catgagggag caaagagctt gaggaatgcc atactccggc 4800 tggtccggga catggaaatt cggactcagg gaggacccgg gctgggcaat gactgggaga 4860 cttgcctggg ttcccaggac ttgggggtcc tgactcccag ccctcatcct gccttacccc 4920 tctgttccca gccccagcct ttctaagcca ttgggaatag aatggcccct tttgttctgg 4980 tgtccagggg tgattgtgcc aaagctctta tttccagtgc caagccccca gaggcttgta 5040 agagttggga tgagggatgg agagggactg ggtctctggg aacaggttgg aggtcttatc 5100 tgtggactgt ctgactccca gctgaggcca agatggggca tgtccccgtc tctgcttagc 5160 gtctgggtga gaaaaacagg ctgtgatcca gaagaaggga agatagagaa ggagggaaag 5220 gatgtaggcg aaggaggtga gagacaggat aggaggaagg aagtggagga ggaggtggta 5280 ggaattggaa ggaggtagaa gccgtgcaga ggaagagggg agagggacga aggaggagcg 5340 atgaagaaga ggagggagac aaaaaaaggg aag 5373

Claims (84)

What is claimed is:

1. An isolated polypeptide selected from the group consisting of:
a) a polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, b) a polypeptide comprising a naturally occurring amino acid sequence at least 90% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, c) a biologically active fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, and d) an immunogenic fragment of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-20.

2. An isolated polypeptide of claim 1 selected from the group consisting of SEQ ID NO:1-20.

3. An isolated polynucleotide encoding a polypeptide of claim 1.

4. An isolated polynucleotide encoding a polypeptide of claim 2.

5. An isolated polynucleotide of claim 4 selected from the group consisting of SEQ ID
NO:21-40.

6. A recombinant polynucleotide comprising a promoter sequence operably linked to a polynucleotide of claim 3.

7. A cell transformed with a recombinant polynucleotide of claim 6.

8. A transgenic organism comprising a recombinant polynucleotide of claim 6.

9. A method for producing a polypeptide of claim 1, the method comprising:
a) culturing a cell under conditions suitable for expression of the polypeptide, wherein said cell is transformed with a recombinant polynucleotide, and said recombinant polynucleotide comprises a promoter sequence operably linked to a polynucleotide encoding the polypeptide of claim 1, and b) recovering the polypeptide so expressed.

10. An isolated antibody which specifically binds to a polypeptide of claim 1.

11. An isolated polynucleotide selected from the group consisting of:
a) a polynucleotide comprising a polynucleotide sequence selected from the group consisting of SEQ ID NO:21-40, b) a polynucleotide comprising a naturally occurring polynucleotide sequence at least 90%
identical to a polynucleotide sequence selected from the group consisting of SEQ ID NO:21-40, c) a polynucleotide complementary to a polynucleotide of a), d) a polynucleotide complementary to a polynucleotide of b), and e) an RNA equivalent of a)-d).

12. An isolated polynucleotide comprising at least 60 contiguous nucleotides of a polynucleotide of claim 11.

13. A method for detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide of claim 11, the method comprising:
a) hybridizing the sample with a probe comprising at least 20 contiguous nucleotides comprising a sequence complementary to said target polynucleotide in the sample, and which probe specifically hybridizes to said target polynucleotide, under conditions whereby a hybridization complex is formed between said probe and said target polynucleotide or fragments thereof, and b) detecting the presence or absence of said hybridization complex, and, optionally, if present, the amount thereof.

14. A method of claim 13, wherein the probe comprises at least 60 contiguous nucleotides.

15. A method for detecting a target polynucleotide in a sample, said target polynucleotide having a sequence of a polynucleotide of claim 11, the method comprising:
a) amplifying said target polynucleotide or fragment thereof using polymerase chain reaction amplification, and b) detecting the presence or absence of said amplified target polynucleotide or fragment thereof, and, optionally, if present, the amount thereof.

16. A composition comprising a polypeptide of claim 1 and a pharmaceutically acceptable excipient.

17. A composition of claim 16, wherein the polypeptide has an amino acid sequence selected from the group consisting of SEQ ID NO:1-20.

18. A method for treating a disease or condition associated with decreased expression of functional PKIN, comprising administering to a patient in need of such treatment the composition of claim 16.

19. A method for screening a compound for effectiveness as an agonist of a polypeptide of claim 1, the method comprising:
a) exposing a sample comprising a polypeptide of claim 1 to a compound, and b) detecting agonist activity in the sample.

20. A composition comprising an agonist compound identified by a method of claim 19 and a pharmaceutically acceptable excipient.

21. A method for treating a disease or condition associated with decreased expression of functional PKIN, comprising administering to a patient in need of such treatment a composition of claim 20.

22. A method for screening a compound for effectiveness as an antagonist of a polypeptide of claim 1, the method comprising:
a) exposing a sample comprising a polypeptide of claim 1 to a compound, and b) detecting antagonist activity in the sample.

23. A composition comprising an antagonist compound identified by a method of claim 22 and a pharmaceutically acceptable excipient.

24. A method for treating a disease or condition associated with overexpression of functional PKIN, comprising administering to a patient in need of such treatment a composition of claim 23.

25. A method of screening for a compound that specifically binds to the polypeptide of claim 1, said method comprising the steps of:
a) combining the polypeptide of claim 1 with at least one test compound under suitable conditions, and b) detecting binding of the polypeptide of claim 1 to the test compound, thereby identifying a compound that specifically binds to the polypeptide of claim 1.

26. A method of screening for a compound that modulates the activity of the polypeptide of claim 1, said method comprising:
a) combining the polypeptide of claim 1 with at least one test compound under conditions permissive for the activity of the polypeptide of claim 1, b) assessing the activity of the polypeptide of claim 1 in the presence of the test compound, and c) comparing the activity of the polypeptide of claim 1 in the presence of the test compound with the activity of the polypeptide of claim 1 in the absence of the test compound, wherein a change in the activity of the polypeptide of claim 1 in the presence of the test compound is indicative of a compound that modulates the activity of the polypeptide of claim 1.

27. A method for screening a compound for effectiveness in altering expression of a target polynucleotide, wherein said target polynucleotide comprises a sequence of claim 5, the method comprising:
a) exposing a sample comprising the target polynucleotide to a compound, under conditions suitable for the expression of the target polynucleotide, b) detecting altered expression of the target polynucleotide, and c) comparing the expression of the target polynucleotide in the presence of varying amounts of the compound and in the absence of the compound.

28. A method for assessing toxicity of a test compound, said method comprising:
a) treating a biological sample containing nucleic acids with the test compound;
b) hybridizing the nucleic acids of the treated biological sample with a probe comprising at least 20 contiguous nucleotides of a polynucleotide of claim 11 under conditions whereby a specific hybridization complex is formed between said probe and a target polynucleotide in the biological sample, said target polynucleotide comprising a polynucleotide sequence of a polynucleotide of claim 11 or fragment thereof;

c) quantifying the amount of hybridization complex; and d) comparing the amount of hybridization complex in the treated biological sample with the amount of hybridization complex in an untreated biological sample, wherein a difference in the amount of hybridization complex in the treated biological sample is indicative of toxicity of the test compound.

29. A diagnostic test for a condition or disease associated with the expression of PKIN in a biological sample comprising the steps of:
a) combining the biological sample with an antibody of claim 10, under conditions suitable for the antibody to bind the polypeptide and form an antibody:polypeptide complex;
and b) detecting the complex, wherein the presence of the complex correlates with the presence of the polypeptide in the biological sample.

30. The antibody of claim 10, wherein the antibody is:
a) a chimeric antibody, b) a single chain antibody, c) a Fab fragment, d) a F(ab')2 fragment, or e) a humanized antibody.

31. A composition comprising an antibody of claim 10 and an acceptable excipient.

32. A method of diagnosing a condition or disease associated with the expression of PKIN in a subject, comprising administering to said subject an effective amount of the composition of claim 31.

33. A composition of claim 31, wherein the antibody is labeled.

34. A method of diagnosing a condition or disease associated with the expression of PKIN in a subject, comprising administering to said subject an effective amount of the composition of claim 33.

35. A method of preparing a polyclonal antibody with the specificity of the antibody of claim comprising:
a) immunizing an animal with a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, or an immunogenic fragment thereof, under conditions to elicit an antibody response;
b) isolating antibodies from said animal; and c) screening the isolated antibodies with the polypeptide, thereby identifying a polyclonal antibody which binds specifically to a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-20.

36. An antibody produced by a method of claim 35.

37. A composition comprising the antibody of claim 36 and a suitable carrier.

38. A method of making a monoclonal antibody with the specificity of the antibody of claim comprising:
a) immunizing an animal with a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-20, or an immunogenic fragment thereof, under conditions to elicit an antibody response;
b) isolating antibody producing cells from the animal;
c) fusing the antibody producing cells with immortalized cells to form monoclonal antibody-producing hybridoma cells;
d) culturing the hybridoma cells; and e) isolating from the culture monoclonal antibody which binds specifically to a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID
NO:1-20.

39. A monoclonal antibody produced by a method of claim 38.

40. A composition comprising the antibody of claim 39 and a suitable carrier.

41. The antibody of claim 10, wherein the antibody is produced by screening a Fab expression library.

42. The antibody of claim 10, wherein the antibody is produced by screening a recombinant immunoglobulin library.

43. A method for detecting a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-20 in a sample, comprising the steps of:
a) incubating the antibody of claim 10 with a sample under conditions to allow specific binding of the antibody and the polypeptide; and b) detecting specific binding, wherein specific binding indicates the presence of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID
NO:1-20 in the sample.

44. A method of purifying a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-20 from a sample, the method comprising:
a) incubating the antibody of claim 10 with a sample under conditions to allow specific binding of the antibody and the polypeptide; and b) separating the antibody from the sample and obtaining the purified polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1-20.

45. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:1.

46. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:2.

47. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:3.

48. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:4.

49. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:5.

50. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:6.

51. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:7.

52. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:8.

53. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:9.

54. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:10.

55. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:11.

56. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:12.

57. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:13.

58. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:14.

59. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:15.

60. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:16.

61. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:17.

62. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:18.

63. A polypeptide of claim l, comprising the amino acid sequence of SEQ ID
NO:19.

64. A polypeptide of claim 1, comprising the amino acid sequence of SEQ ID
NO:20.

65. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:21.

66. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:22.

67. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:23.

68. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:24.

69. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:25.

70. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:26.

71. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:27.

72. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:28.

73. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:29.

74. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:30.

75. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:31.

76. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:32.

77. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:33.

78. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:34.

79. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:35.

80. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:36.

81. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:37.

82. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:38.

83. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:39.

84. A polynucleotide of claim 11, comprising the polynucleotide sequence of SEQ ID NO:40.