CN1155614C - Human protein with cancer cell growth suppressing function and its coding sequence - Google Patents

Abstract

The present invention discloses a novel human protein with the function of inhibiting cancer, polynucleotide for encoding the polypeptide and a method for preparing the polypeptide by a recombinant technology. The present invention also discloses a method of using the polypeptide to treat various diseases, such as cancers. The present invention also discloses an antagonist of the polypeptide and a therapeutic effect thereof. The present invention also discloses the application of the polynucleotide for encoding the human protein with the function of inhibiting cancer.

Description

New people's albumen and encoding sequence thereof with anticancer growth function

The invention belongs to biological technical field, specifically, the present invention relates to the proteic polynucleotide of people that new coding has cancer suppressing function, and the polypeptide of this polynucleotide encoding.The invention still further relates to the purposes and the preparation of these polynucleotide and polypeptide.

The research of people's gene group is international focus at present, removes human chromosome DNA large scale sequencing, outside the method for expressed sequence order-checking (EST), also lacks the screening that begins from function and has the high-throughout method of functional gene.

Cancer is one of principal disease of harm humans health.In order to treat effectively and prophylaxis of tumours, people more and more pay close attention to genetic treatment of tumor at present.Therefore, this area presses for people's albumen and the agonist/inhibitor thereof that development research has cancer suppressing function.

The purpose of this invention is to provide the new people's protein polypeptide of a class with cancer suppressing function with and fragment, analogue and derivative.

Another object of the present invention provides the polynucleotide of these polypeptide of coding.

Another object of the present invention provides the method for these polypeptide of production and the purposes of this polypeptide and encoding sequence.

In a first aspect of the present invention, novel isolated protein polypeptide with cancer suppressing function is provided, and it comprises the polypeptide of the aminoacid sequence with the group of being selected from down: SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ IDNO:11, SEQ ID NO:14, SEQ ID NO:17, SEQ ID NO:20; Or its conservative property variation polypeptide or its active fragments or its reactive derivative.

Preferably, this polypeptide is the polypeptide with aminoacid sequence of the group of being selected from down: SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:11, SEQ ID NO:14, SEQ ID NO:17, SEQ ID NO:20.

In a second aspect of the present invention, a kind of isolating polynucleotide are provided, it comprises a nucleotide sequence, and this nucleotide sequence is shown at least 85% homogeny with a kind of nucleotides sequence that is selected from down group: the polynucleotide of the above-mentioned protein polypeptide with cancer suppressing function of (a) encoding; (b) with polynucleotide (a) complementary polynucleotide.Preferably, the polypeptide of this polynucleotide encoding has the aminoacid sequence of the group of being selected from down: SEQ D NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ IDNO:11, SEQ ID NO:14, SEQ ID NO:17, SEQ ID NO:20.More preferably, the sequence of these polynucleotide is selected from down group: coding region sequence or the full length sequence of SEQ ID NO:3, SEQ ID NO:6, SEQ ID NO:9, SEQ ID NO:12, SEQ ID NO:15, SEQ ID NO:18, SEQ ID NO:21.

In a third aspect of the present invention, the carrier that contains above-mentioned polynucleotide is provided, and has been transformed or host cell of transduceing or the host cell that is directly transformed or transduce by above-mentioned polynucleotide by this carrier.

In a fourth aspect of the present invention, the preparation method who prepares the polypeptide of the protein-active with cancer suppressing function is provided, this method comprises: (a) have under the proteic condition of cancer suppressing function suitable the expression, cultivate the above-mentioned host cell that is transformed or transduce; (b) from culture, isolate the polypeptide of protein-active with cancer suppressing function.

In a fifth aspect of the present invention, provide and above-mentioned protein polypeptide specificity bonded antibody with cancer suppressing function.The nucleic acid molecule that can be used for detecting also is provided, and it contains, and continuous 10 Nucleotide are to full length nucleotide in the above-mentioned polynucleotide, and preferably it contains the about 10-800 of a successive Nucleotide.

In a sixth aspect of the present invention, a kind of pharmaceutical composition is provided, it contains the protein polypeptide and the pharmaceutically acceptable carrier with cancer suppressing function of the present invention of safe and effective amount.These pharmaceutical compositions can be treated illnesss such as cancer and cellular abnormality propagation.

Others of the present invention are because disclosing of the technology of this paper is conspicuous to those skilled in the art.

The present invention adopts large-scale cDNA clone transfection cancer cells, has on the basis of cancer suppressing action in acquisition, proves new gene through order-checking, further obtains full length cDNA clone.DNA transfection evidence, the albumen with cancer suppressing function of the present invention has the effect that suppresses clone's formation to cancer cells (liver cancer cell), and its inhibiting rate is more than 50% or 50%.

As used herein, " isolating " is meant that material separates (if natural substance, primal environment promptly is a natural surroundings) from its primal environment.Do not have separation and purification as polynucleotide under the native state in the active somatic cell and polypeptide, but same polynucleotide or polypeptide as from native state with in other materials that exist separately, then for separation and purification.

As used herein, " isolating albumen or polypeptide with cancer suppressing function " is meant that the protein polypeptide with cancer suppressing function is substantially free of natural relative other albumen, lipid, carbohydrate or other material.Those skilled in the art can have the albumen of cancer suppressing function with the purified technology of protein purifying of standard.Basically pure polypeptide can produce single master tape on non-reduced polyacrylamide gel.Purity with protein polypeptide of cancer suppressing function can be used amino acid sequence analysis.

Polypeptide of the present invention can be recombinant polypeptide, natural polypeptides, synthetic polypeptide, preferred recombinant polypeptide.Polypeptide of the present invention can be the product of natural purifying, or the product of chemosynthesis, or uses recombinant technology to produce from protokaryon or eucaryon host (for example, bacterium, yeast, higher plant, insect and mammalian cell).The host used according to the recombinant production scheme, polypeptide of the present invention can be glycosylated, maybe can be nonglycosylated.Polypeptide of the present invention also can comprise or not comprise initial methionine residues.

The present invention also comprises the proteic fragment of the people with cancer suppressing function, derivative and analogue.As used herein, term " fragment ", " derivative " are meant with " analogue " and keep natural identical biological function or the active polypeptide of people's albumen with cancer suppressing function of the present invention basically.Polypeptide fragment of the present invention, derivative or analogue can be that (i) has one or more conservative or substituted polypeptide of non-conservation amino-acid residue (preferred conservative amino acid residue), and the amino-acid residue of such replacement can be also can not encoded by genetic code, or (ii) in one or more amino-acid residues, has a polypeptide of substituted radical, or (iii) mature polypeptide and another compound (such as the compound that prolongs the polypeptide transformation period, polyoxyethylene glycol for example) merge formed polypeptide, or (iv) additional aminoacid sequence is fused to this peptide sequence and the polypeptide that forms (as leader sequence or secretion sequence or be used for the sequence or the proteinogen sequence of this polypeptide of purifying).According to the instruction of this paper, these fragments, derivative and analogue belong to the known scope of those skilled in the art.

Polynucleotide of the present invention can be dna form or rna form.Dna form comprises the DNA of cDNA, genomic dna or synthetic.DNA can be strand or double-stranded.DNA can be coding strand or noncoding strand.Be example with PP8153 albumen (in this application, its clone numbering is adopted in proteinic name), the coding region sequence of encoding mature polypeptide can be identical with the coding region sequence shown in the SEQ ID NO:3 or the varient of degeneracy.As used herein, " varient of degeneracy " is meant that in the present invention coding has the protein of SEQ ID NO:2, but with the differentiated nucleotide sequence of coding region sequence shown in the SEQ IDNO:3.Be example with PP8332 albumen (in this application, its clone numbering is adopted in proteinic name), the coding region sequence of encoding mature polypeptide can be identical with the coding region sequence shown in the SEQ ID NO:6 or the varient of degeneracy.As used herein, " varient of degeneracy " is meant that in the present invention coding has the protein of SEQ ID NO:5, but with the differentiated nucleotide sequence of coding region sequence shown in the SEQ ID NO:6.Have the albumen of cancer suppressing function for other, can the rest may be inferred.Have the albumen of cancer suppressing function for other, can the rest may be inferred.

The polynucleotide of encoding mature polypeptide comprise: the encoding sequence of an encoding mature polypeptide; The encoding sequence of mature polypeptide and various additional code sequence; Encoding sequence of mature polypeptide (with optional additional code sequence) and non-coding sequence.

Term " polynucleotide of coded polypeptide " can be the polynucleotide that comprise this polypeptide of encoding, and also can be the polynucleotide that also comprise additional code and/or non-coding sequence.

The invention still further relates to the varient of above-mentioned polynucleotide, its coding has the polypeptide of identical aminoacid sequence or fragment, analogue and the derivative of polypeptide with the present invention.The varient of these polynucleotide can be the allelic variant of natural generation or the varient that non-natural takes place.These nucleotide diversity bodies comprise and replace varient, deletion mutation body and insert varient.As known in the art, allelic variant is the replacement form of polynucleotide, and it may be replacement, disappearance or the insertion of one or more Nucleotide, but can be from not changing the function of its encoded polypeptides in fact.

The invention still further relates to and above-mentioned sequence hybridization and two sequences between have at least 50%, preferably at least 70%, the polynucleotide of at least 80% homogeny more preferably.The present invention be more particularly directed under stringent condition and the interfertile polynucleotide of polynucleotide of the present invention.In the present invention, " stringent condition " is meant: (1) than hybridization under low ionic strength and the comparatively high temps and wash-out, as 0.2 * SSC, and 0.1%SDS, 60 ℃; Or (2) hybridization the time is added with denaturing agent, as 50% (v/v) methane amide, 0.1% calf serum/0.1%Ficoll, 42 ℃ etc.; Or (3) only at the homogeny between the two sequences at least more than 95%, be more preferably 97% and just hybridize when above.And the polypeptide of interfertile polynucleotide encoding has identical biological function (is example with PP8153 albumen) and activity with the mature polypeptide shown in the SEQ IDNO:2.

The invention still further relates to nucleic acid fragment with above-mentioned sequence hybridization.As used herein, the length of " nucleic acid fragment " contains 15 Nucleotide at least, better is at least 30 Nucleotide, is more preferably at least 50 Nucleotide, preferably more than at least 100 Nucleotide.The amplification technique (as PCR) that nucleic acid fragment can be used for nucleic acid has the proteic polynucleotide of cancer suppressing function to determine and/or to separate to encode.

Polypeptide among the present invention and polynucleotide preferably provide with isolating form, more preferably are purified to homogeneous.

Dna sequence dna of the present invention can obtain with several method.For example, with hybridization technique DNA isolation well known in the art.These technology including, but not limited to: 1) with probe and genome or the hybridization of cDNA library to detect homology nucleotide sequence and 2) antibody screening of expression library to be to detect the dna fragmentation of the clone with common structure feature.

The proteic specific DNA fragment sequence that coding has cancer suppressing function produces also and can obtain with following method: 1) separate double chain DNA sequence from genomic dna; 2) the chemical synthesising DNA sequence is to obtain the double-stranded DNA of required polypeptide.

In the above-mentioned method of mentioning, isolation of genomic DNA is least commonly used.When the whole aminoacid sequence of the polypeptide product of needs was known, the direct chemical of dna sequence dna is synthetic to be the method for often selecting for use.When if required amino acid whose whole sequence is not known, the direct chemical of dna sequence dna is synthetic to be impossible, and the method for selecting for use is the separation of cDNA sequence.The standard method that separates interested cDNA is from the donorcells separating mRNA of this gene of high expression level and carries out reverse transcription, forms plasmid or phage cDNA library.Extract the existing multiple proven technique of method of mRNA, test kit also can obtain (Qiagene) from commercial channels.And the construction cDNA library also is usual method (Sambrook, et al., Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory.New York, 1989).Also can obtain the cDNA library of commercial offers, as the different cDNA library of Clontech company.When being used in combination the polymeric enzyme reaction technology, even few expression product also can be cloned.

Available ordinary method is screened gene of the present invention from these cDNA libraries.These methods include, but is not limited to: (1) DNA-DNA or DNA-RNA hybridization; (2) function of marker gene occurs or forfeiture; (3) mensuration has the level of the proteic transcript of cancer suppressing function; (4), detect the protein product of genetic expression by immunological technique or mensuration biologic activity.Aforesaid method can singly be used, but also several different methods combined utilization.

In (1) kind method, hybridizing used probe is and any a part of homology of polynucleotide of the present invention that at least 15 Nucleotide of its length better are at least 30 Nucleotide, are more preferably at least 50 Nucleotide, preferably at least 100 Nucleotide.In addition, the length of probe within 2kb, preferably is within the 1kb usually.Probe used herein is the dna sequence dna of chemosynthesis on the basis of gene DNA sequence information of the present invention normally.Gene of the present invention itself or fragment are certainly as probe.The mark of dna probe can be used radio isotope, fluorescein or enzyme (as alkaline phosphatase) etc.

In (4) kind method, detect the protein product of protein gene expression and can use immunological technique such as Western blotting, radioimmunoprecipitation, enzyme-linked immunosorbent assay (ELISA) etc. with cancer suppressing function.

Use method (Saiki, the et al.Science 1985 of round pcr DNA amplification/RNA; 230:1350-1354) be optimized for acquisition gene of the present invention.When particularly being difficult to obtain the cDNA of total length from the library, can preferably use RACE method (the terminal rapid amplifying method of RACE-cDNA), the primer that is used for PCR can suitably be selected according to sequence information of the present invention disclosed herein, and available ordinary method is synthetic.Available ordinary method is as the DNA/RNA fragment by gel electrophoresis separation and purifying amplification.

The gene of the present invention that obtains as mentioned above, perhaps the available ordinary method of mensuration of the nucleotide sequence of various dna fragmentations etc. such as dideoxy chain termination (Sanger et al.PNAS, 1977,74:5463-5467).This class nucleotide sequencing is available commercial sequencing kit etc. also.In order to obtain the cDNA sequence of total length, order-checking need be carried out repeatedly.Sometimes need to measure a plurality of clones' cDNA sequence, just can be spliced into the cDNA sequence of total length.

The present invention also relates to comprise the carrier of polynucleotide of the present invention, and the host cell that produces through genetically engineered with carrier of the present invention or albumen coded sequence with cancer suppressing function, and the method that produces polypeptide of the present invention through recombinant technology.

Recombinant DNA technology (Science, 1984 by routine; 224:1431), can utilize polymerized nucleoside acid sequence of the present invention to can be used to express or produce the protein polypeptide with cancer suppressing function of reorganization.In general following steps are arranged:

(1). have the proteic polynucleotide of people (or varient) of cancer suppressing function with coding of the present invention, or transform or the transduction proper host cell with the recombinant expression vector that contains these polynucleotide;

(2). the host cell of in suitable medium, cultivating;

(3). separation, protein purification from substratum or cell.

Among the present invention, the people's albumen polynucleotide sequence with cancer suppressing function can be inserted in the recombinant expression vector.Term " recombinant expression vector " refers to that bacterial plasmid well known in the art, phage, yeast plasmid, vegetable cell virus, mammalian cell virus are as adenovirus, retrovirus or other carriers.The carrier of Shi Yonging includes but not limited in the present invention: and the expression vector based on T7 of in bacterium, expressing (Rosenberg, et al.Gene, 1987,56:125); The pMSXND expression vector of in mammalian cell, expressing (Lee and Nathans, J Bio Chem.263:3521,1988) and at the carrier that derives from baculovirus of expressed in insect cells.In a word, as long as can duplicate in host and stablize, any plasmid and carrier can be used.A key character of expression vector is to contain replication orgin, promotor, marker gene and translation controlling elements usually.

Method well-known to those having ordinary skill in the art can be used to make up and contains people's encoding histone dna sequence dna with cancer suppressing function and suitable transcribing/the translate expression vector of control signal.These methods comprise (Sambroook, et al.Molecular Cloning, a Laboratory Manual, coldSpring Harbor Laboratory.New York, 1989) such as extracorporeal recombinant DNA technology, DNA synthetic technology, the interior recombinant technologys of body.Described dna sequence dna can effectively be connected on the suitable promotor in the expression vector, and is synthetic to instruct mRNA.The representative example of these promotors has: colibacillary lac or trp promotor; Lambda particles phage P _LPromotor; Eukaryotic promoter comprises LTRs and some other known may command gene expression promoter in protokaryon or eukaryotic cell or its virus of CMV immediate early promoter, HSV thymidine kinase promoter, early stage and late period SV40 promotor, retrovirus.Expression vector also comprises ribosome bind site and the transcription terminator that translation initiation is used.

In addition, expression vector preferably comprises one or more selected markers, to be provided for selecting the phenotypic character of transformed host cells, cultivate Tetrahydrofolate dehydrogenase, neomycin resistance and the green fluorescent protein (GFP) of usefulness as eukaryotic cell, or be used for colibacillary tsiklomitsin or amicillin resistance.

Comprise the carrier of above-mentioned suitable dna sequence dna and suitable promotor or control sequence, can be used to transform appropriate host cell, so that it can marking protein.

Host cell can be a prokaryotic cell prokaryocyte, as bacterial cell; Or eukaryotic cell such as low, as yeast cell; Or higher eucaryotic cells, as mammalian cell.Representative example has: intestinal bacteria, streptomyces; The bacterial cell of Salmonella typhimurium; Fungal cell such as yeast; Vegetable cell; The insect cell of fruit bat S2 or Sf9; The zooblast of CHO, COS or Bowes melanoma cells etc.

When polynucleotide of the present invention are expressed in higher eucaryotic cells, be enhanced if will make to transcribe when in carrier, inserting enhancer sequence.Enhanser is the cis acting factor of DNA, and nearly 10 to 300 base pairs act on promotor transcribing with enhancing gene usually.Can for example be included in the SV40 enhanser of 100 to 270 base pairs of replication origin side in late period one, at the polyoma enhanser of replication origin side in late period one and adenovirus enhanser etc.

Persons skilled in the art all know how to select appropriate carriers, promotor, enhanser and host cell.

Can carry out with routine techniques well known to those skilled in the art with the recombinant DNA transformed host cell.When the host was prokaryotic organism such as intestinal bacteria, the competent cell that can absorb DNA can be used CaCl in exponential growth after date results ₂Method is handled, and used step is well-known in this area.Alternative is to use MgCl ₂If desired, transforming also the method for available electroporation carries out.When the host is an eukaryote, can select following DNA transfection method for use: coprecipitation of calcium phosphate method, conventional mechanical method such as microinjection, electroporation, liposome packing etc.

The transformant that obtains can be cultivated with ordinary method, expresses the polypeptide of coded by said gene of the present invention.According to used host cell, used substratum can be selected from various conventional substratum in the cultivation.Under the condition that is suitable for the host cell growth, cultivate.After host cell grows into suitable cell density, induce the promotor of selection with suitable method (as temperature transition or chemical induction), cell is cultivated for some time again.

Recombinant polypeptide in the above methods can wrap by in cell, extracellular or on cytolemma, express or be secreted into the extracellular.If desired, can utilize its physics, the separating by various separation methods with other characteristic and the albumen of purification of Recombinant of chemistry.These methods are well-known to those skilled in the art.The example of these methods includes, but are not limited to: conventional renaturation handles, with protein precipitant handle (salt analysis method), centrifugal, the broken bacterium of infiltration, superly handle, the combination of super centrifugal, sieve chromatography (gel-filtration), adsorption chromatography, ion exchange chromatography, high performance liquid chromatography (HPLC) and other various liquid chromatography (LC) technology and these methods.

The people's albumen or the polypeptide with cancer suppressing function of reorganization are of use in many ways.These purposes include, but is not limited to: directly have the disease due to the low or forfeiture of the protein function of cancer suppressing function as pharmacological agent and be used to screen and promote or antagonism has antibody, polypeptide or other part of the protein function of cancer suppressing function.For example, antibody can be used for activating or suppressing to have the proteic function of people of cancer suppressing function.The people's protein screening peptide library that has a cancer suppressing function with the reorganization of expressing can be used for seeking the peptide molecule that can suppress or stimulate the people's protein function with cancer suppressing function of therapeutic value.

The present invention also provides screening of medicaments to improve (agonist) or check the method that (antagonist) has the proteic medicament of people of cancer suppressing function to identify.Agonist improves the biological function such as stimulate cellular proliferation of the people's albumen with cancer suppressing function, and antagonist prevention disorder such as the various cancer relevant with cell hyperproliferation with treatment.For example, can be in the presence of medicine, the proteic film preparation of people that mammalian cell or expression is had cancer suppressing function is cultivated with the people's albumen with cancer suppressing function of mark.Measure the medicine raising then or check this interactional ability.

The proteic antagonist of people with cancer suppressing function comprises antibody, compound, acceptor disappearance thing and the analogue etc. that filter out.The proteic antagonist of people with cancer suppressing function can and be eliminated its function with the people's protein binding with cancer suppressing function, or suppresses to have the proteic generation of people of cancer suppressing function, or combines with the avtive spot of polypeptide and to make polypeptide can not bring into play biological function.The proteic antagonist of people with cancer suppressing function can be used for therepic use.

In screening during as the compound of antagonist, the albumen that can have a cancer suppressing function adds during bioanalysis measures, and determines by measuring albumen and the interaction between its acceptor that compounds affect has cancer suppressing function whether compound is antagonist.With the same quadrat method of above-mentioned SCREENED COMPOUND, can filter out the acceptor disappearance thing and the analogue of antagonist action.

Polypeptide of the present invention can be directly used in disease treatment, for example, and various malignant tumours and cellular abnormality propagation etc.

Polypeptide of the present invention, and fragment, derivative, analogue or their cell can be used as antigen to produce antibody.These antibody can be polyclone or monoclonal antibody.Polyclonal antibody can obtain by the method with this polypeptide direct injection animal.The technology of preparation monoclonal antibody comprises hybridoma technology, three knurl technology, people B-quadroma technology, EBV-hybridoma technology etc.

Can be with polypeptide of the present invention and antagonist and suitable pharmaceutical carrier combination back use.These carriers can be water, glucose, ethanol, salt, damping fluid, glycerine and their combination.Composition comprises the polypeptide or the antagonist of safe and effective amount and carrier and the vehicle that does not influence effect of drugs.These compositions can be used as medicine and are used for disease treatment.

The present invention also provides medicine box or the test kit that contains one or more containers, and one or more medicinal compositions compositions of the present invention are housed in the container.With these containers, can have by the given indicative prompting of government authorities of making, using or selling medicine or biological products, the government authorities that this prompting reflects production, uses or sells permits it to use on human body.In addition, polypeptide of the present invention can be used in combination with other treatment compound.

Pharmaceutical composition can be with mode administration easily, as by in part, intravenously, intraperitoneal, intramuscular, subcutaneous, the nose or the route of administration of intracutaneous.Albumen with cancer suppressing function comes administration with the amount that treats and/or prevents concrete indication effectively.The proteic amount with cancer suppressing function and the dosage range that are applied to the patient will depend on many factors, as administering mode, person's to be treated healthiness condition and diagnostician's judgement.

The proteic polynucleotide of people with cancer suppressing function also can be used for multiple therapeutic purpose.Gene therapy technology can be used for treating since have that the proteic nothing of cancer suppressing function is expressed or the proteic expression with cancer suppressing function of unusual/non-activity due to cell proliferation, growth or metabolic disturbance.The albumen with cancer suppressing function that the gene therapy vector (as virus vector) of reorganization can be designed to express variation is to suppress endogenic protein-active with cancer suppressing function.For example, a kind of albumen with cancer suppressing function of variation can be the albumen with cancer suppressing function that shortens, lacked signal conduction function territory, though can combine with the substrate in downstream, lacks signaling activity.Therefore the gene therapy vector of reorganization can be used for treating the protein expression with cancer suppressing function or the disease of active caused by abnormal.Deriving from the expression vector of virus such as protein gene that retrovirus, adenovirus, adeno-associated virus (AAV), hsv, parvovirus etc. can be used for having cancer suppressing function is transferred in the cell.The method that structure carries the recombinant viral vector of the protein gene with cancer suppressing function is found in existing document (Sambrook, et al.).The people protein gene of reorganization with cancer suppressing function can be packaged in the liposome and be transferred in the cell in addition.

Suppress to have cancer suppressing function people's protein mRNA oligonucleotide (comprising sense-rna and DNA) and ribozyme also within the scope of the invention.Ribozyme is the enzyme sample RNA molecule that a kind of energy specificity is decomposed specific RNA, and its mechanism of action is to carry out the endonuclease effect after ribozyme molecule and the hybridization of complementary target RNA-specific.The RNA of antisense and DNA and ribozyme can obtain with existing any RNA or DNA synthetic technology, as the technology widespread use of solid phase phosphoamide chemical synthesis synthetic oligonucleotide.Antisense rna molecule can be transcribed acquisition by the dna sequence dna of this RNA that encodes in external or body.This dna sequence dna has been incorporated into the downstream of rna polymerase promoter of carrier.In order to increase the stability of nucleic acid molecule, available several different methods is modified it, and as increasing the sequence length of both sides, the connection between the ribonucleoside is used phosphoric acid thioester bond or peptide bond but not phosphodiester bond.

Polynucleotide imports tissue or intracellular method comprises: directly be injected into polynucleotide in the in-vivo tissue; Or external by carrier (as virus, phage or plasmid etc.) earlier with the polynucleotide transfered cell in, again cell is transplanted in the body etc.

Polypeptide of the present invention also can be used as the peptide spectrum analysis, for example, the polypeptide available physical, chemistry or enzyme carry out the specificity cutting, and carry out the two-dimentional or three-dimensional gel electrophoresis analysis of one dimension.

The present invention also provides the antibody at the people's proteantigen determinant with cancer suppressing function.These antibody include, but is not limited to: the fragment that polyclonal antibody, monoclonal antibody, chimeric antibody, single-chain antibody, Fab fragment and Fab expression library produce.

The anti-proteic antibody of people with cancer suppressing function can be used in the immunohistochemistry technology, detects the people's albumen with cancer suppressing function in the biopsy specimen.

With the also available labelled with radioisotope of the protein bound monoclonal antibody of the people with cancer suppressing function, inject in the body and can follow the tracks of its position and distribution.This radiolabeled antibody can be used as a kind of atraumatic diagnostic method and is used for the location of tumour cell and has judged whether transfer.

Antibody among the present invention can be used for treating or prevents and the relevant disease of people's albumen with cancer suppressing function.The antibody that gives suitable dosage can stimulate or block proteic generation of the people with cancer suppressing function or activity.

Antibody also can be used for designing the immunotoxin at a certain privileged sites in the body.As have cancer suppressing function people's albumen high-affinity monoclonal antibody can with bacterium or plant poison (as diphtheria toxin, ricin, abrine etc.) covalent attachment.A kind of usual method is with sulfydryl linking agent such as SPDP, attacks the amino of antibody, by the exchange of disulfide linkage, toxin is incorporated on the antibody, and this hybrid antibody can be used for killing the cell of the people's protein positive with cancer suppressing function.

Available people's albumen or the polypeptide immune animal of the production of polyclonal antibody with cancer suppressing function, as rabbit, mouse, rat etc.Multiple adjuvant can be used for the enhancing immunity reaction, includes but not limited to freund's adjuvant etc.

Have cancer suppressing function people's protein monoclonal antibody can with hybridoma technology production (Kohler and Milstein.Nature, 1975,256:495-497).With the variable region bonded chimeric antibody in human constant region and inhuman source can with existing technology production (Morrison et al, PNAS, 1985,81:6851).And the technology of existing manufacture order chain antibody (U.S.PatNo.4946778) also can be used for producing the anti-proteic single-chain antibody of people with cancer suppressing function.

Can be incorporated into the rondom polypeptide storehouse that solid formation forms by the various amino acid that may make up by screening with the protein bound peptide molecule of the people with cancer suppressing function obtains.During screening, must carry out mark to people's protein molecular with cancer suppressing function.

The invention still further relates to quantitatively and detection and localization has the diagnostic testing process of people's protein level of cancer suppressing function.These tests are known in the art, and comprise that FISH measures and radioimmunoassay.The people's protein level that is detected in the test with cancer suppressing function, the disease that can have the importance of people's albumen in various diseases of cancer suppressing function with laying down a definition and be used to diagnose albumen to work with cancer suppressing function.

Proteic polynucleotide with cancer suppressing function can be used for having the diagnosis and the treatment of the protein related diseases of cancer suppressing function.Aspect diagnosis, the proteic polynucleotide with cancer suppressing function can be used for detecting have cancer suppressing function proteic expression whether or under morbid state, have an abnormal exprssion of cancer suppressing function.As the protein D NA sequence with cancer suppressing function can be used for the hybridization of biopsy specimen is had with judgement the proteic abnormal expression of cancer suppressing function.Hybridization technique comprises the Southern blotting, Northern blotting, in situ hybridization etc.These technological methods all are disclosed mature technologies, and relevant test kit all can obtain from commercial channels.Part or all of polynucleotide of the present invention can be used as probe stationary on microarray (Microarray) or DNA chip (being called " gene chip " again), is used for analyzing the differential expression analysis and the gene diagnosis of tissue gene.Carry out RNA-polymerase chain reaction (RT-PCR) amplification in vitro with the special primer of the albumen with cancer suppressing function and also can detect proteic transcription product with cancer suppressing function.

The sudden change that detection has the protein gene of cancer suppressing function also can be used for diagnosing the relevant disease of albumen with cancer suppressing function.Form with protein mutation of cancer suppressing function comprises that to have point mutation that the protein D NA sequence of cancer suppressing function compares, transposition, disappearance, reorganization and other any unusual etc. with normal wild type.Available existing technology such as Southern blotting, dna sequence analysis, PCR and in situ hybridization detect sudden change.In addition, sudden change might influence proteic expression, therefore can judge indirectly that with Northern blotting, Western blotting gene has or not sudden change.

Sequence of the present invention identifies it also is valuable to karyomit(e).This sequence can be specifically at certain bar human chromosome particular location and and can with its hybridization.At present, need to identify the concrete site of each gene on the karyomit(e).Now, have only chromosomal marker thing seldom to can be used for the marker chromosomes position based on actual sequence data (repetition polymorphism).According to the present invention, for these sequences are associated with disease related gene, its important the first step is positioned these dna sequence dnas on the karyomit(e) exactly.

In brief, prepare PCR primer (preferred 15-35bp), sequence can be positioned on the karyomit(e) according to eDNA.Then, these primers are used for the somatocyte hybrid cell that the PCR screening contains each bar human chromosome.Have only those hybrid cells that contain corresponding to the people's gene of primer can produce the fragment of amplification.

The PCR localization method of somatocyte hybrid cell is that DNA is navigated to concrete chromosomal quick method.Use Oligonucleolide primers of the present invention,, can utilize one group to realize inferior location from specific chromosomal fragment or a large amount of genomic clone by similar approach.Other the similar strategy that can be used for chromosomal localization comprises in situ hybridization, uses the karyomit(e) prescreen and the hybridization preliminary election of the airflow classification of mark, thereby makes up the special cDNA storehouse of karyomit(e).

The cDNA clone is carried out fluorescence in situ hybridization (FISH) with Metaphase Chromosome, can in a step, accurately carry out chromosomal localization.The summary of this technology is referring to Verma etc., Human Chromosomes:a Manual of BasicTechniques, Pergamon Press, New York (1988).

In case sequence is positioned to chromosome position accurately, the physical location of this sequence on karyomit(e) just can be associated with the gene map data.These data for example are found in, V.Mckusick, Mendelian Inheritance in Man (can by with the online acquisition of Johns Hopkins University Welch Medical Library).Can pass through linkage analysis then, determine gene and navigated to relation between the disease on the chromosomal region already.

Then, need to measure ill and not cDNA between diseased individuals or genome sequence difference.If observe certain sudden change in some or all of diseased individuals, and this sudden change is not observed in any normal individual, then this sudden change may be the cause of disease of disease.More ill and diseased individuals not is usually directed at first seek the variation of structure in the karyomit(e), as from the horizontal visible of karyomit(e) or use based on detectable disappearance of the PCR of cDNA sequence or transposition.Resolving power according to present physical mapping and assignment of genes gene mapping technology, being accurately positioned to the cDNA of the chromosomal region relevant with disease, can be a kind of (the supposing that 1 megabasse mapping resolving power and every 20kb are corresponding to a gene) between 50 to 500 potential Disease-causing genes.

Pyrenoids thuja acid full length sequence or its fragment with cancer suppressing function of the present invention can obtain with the method for pcr amplification method, recombination method or synthetic usually.For the pcr amplification method, can be disclosed according to the present invention about nucleotide sequence, especially open reading frame sequence designs primer, and with commercially available cDNA storehouse or by the prepared cDNA storehouse of ordinary method well known by persons skilled in the art as template, amplification and must relevant sequence.When sequence is longer, usually needs to carry out twice or pcr amplification repeatedly, and then the fragment that each time amplifies is stitched together by proper order.

In case obtained relevant sequence, just can obtain relevant sequence in large quantity with recombination method.This normally is cloned into carrier with it, changes cell again over to, separates obtaining relevant sequence then from the host cell after the propagation by ordinary method.

In addition, also the method for available synthetic is synthesized relevant sequence, especially fragment length more in short-term.Usually, by first synthetic a plurality of small segments, and then connect and to obtain the very long fragment of sequence.

At present, can be fully come the dna sequence dna of code book invention albumen (or its fragment, or derivatives thereof) by chemosynthesis.This dna sequence dna can be introduced then in the various dna moleculars (as carrier) and cell in this area.In addition, also can will suddenly change and introduce in the protein sequence of the present invention by chemosynthesis.

In addition, because the albumen with cancer suppressing function of the present invention has the natural acid sequence that is derived from the people, therefore, compare with the albumen of the same clan that derives from other species, estimate to have higher active and/or lower side effect (for example in the intravital immunogenicity of people lower or do not have) being applied to man-hour.

Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiment only to be used to the present invention is described and be not used in and limit the scope of the invention.The experimental technique of unreceipted actual conditions in the following example, usually according to people such as normal condition such as Sambrook, molecular cloning: laboratory manual (New York:Cold Spring Harbor LaboratoryPress, 1989) condition described in, or the condition of advising according to manufacturer.

The acquisition of embodiment 1:cDNA gene and the restraining effect that the cancer cells clone is formed

SP2114a come from the liver cDNA library buied from GIBCO BRL company (catalog number (Cat.No.): 10422-012), PP8153, PP8332, PP9177, PP9445, PP10199 and PP10226 obtain by making up the human placenta cDNA library with ordinary method.Get the placenta tissue at 3,6,10 monthly ages, (GIBCOBRL company) extracts total RNA by manufacturer's specification sheets with Trizol reagent, extracts mRNA with the mRNA test kit (Pharmacia company) of purifying.Make up the cDNA library of above-mentioned mRNA with pCMV-script TMXR cDNA library construction test kit (Stratagene company).Wherein ThermoScript II is used MMLV-RT-Superscript II (GIBCO BRL) instead, and reverse transcription reaction carries out at 42 ℃.Transform XL 10-Gold recipient cell, obtained 1 * 10 ⁶The cDNA library of cfu/ μ g cDNA titre.The first round is picking cDNA clone at random, is probe with high abundance cDNA clone with the cDNA clone who has proved cancer inhibitor cell growth function thereafter, screening by hybridization cDNA library, weak positive and negative clone of picking.With Qiagen 96 orifice plate plasmid extraction test kits, carry out the extraction of plasmid DNA by shop instruction.Plasmid DNA and empty carrier transfection simultaneously hepatoma cell line 7721.After the 100ng DNA alcohol precipitation drying, add 6 μ l H ₂Transfection is treated in the O dissolving.Add 0.74 μ l liposome and 9.3 μ l serum-free mediums in every part of DNA sample, behind the mixing, room temperature was placed 10 minutes.Add 150 μ l serum-free mediums in every pipe, divide equally and add 3 holes and grow in 7721 cells of 96 orifice plates, placed 2 hours for 37 ℃, every hole adds 50 μ l serum-free mediums again, 37 ℃ 24 hours.Every hole is changed 100 μ l and is trained liquid entirely, 37 ℃ 24 hours, change the full training liquid 100 μ l that contain G418,37 ℃ 24～48 hours, the limit is observed, the training liquid that G418 concentration does not wait is changed on the limit.After about 2～3 times, there is the clone to form up to the microscopy cell, counting.Find that above clone has the cell clone of inhibition formation effect, the result is as shown in the table.

CDNA clone's transfectional cell (7721) clone formation situation

CDNA clones title	CDNA clones number (three repetitions)	Empty carrier clone number (three repetitions)
			PP8153 PP8332 PP9177 PP9445 PP10199 PP10226 SP2114a	2 0 1 8 3 5 16 11 15 6 1 9 5 2 2 6 1 3 4 6 5	26 29 30 12 13 15 48 38 35 48 38 35 48 38 35 48 38 35 38 42 40

Above-mentioned cDNA clone is adopted two deoxidation cessation method, on the ABI377 automatic dna sequencer, measure the nucleotide sequence of the nearly 500bp of one end.After the analysis, be defined as novel gene cloning, carry out the other end order-checking, do not obtain full length cDNA sequence yet, the design primer checks order once more, up to obtaining full length sequence (SEQ ID NO:1,4,7,10,13,16,19).

Embodiment 2: PCR obtains full-length gene from placenta cDNA:

Get the placenta tissue at 3,6,10 monthly ages, (GIBCO BRL company) extracts total RNA by manufacturer's specification sheets with Trizol reagent, extracts mRNA with the mRNA test kit (Pharmacia company) of purifying.With MMLV-RT-Superscript II (GIBCO BRL), ThermoScript II is carried out reverse transcription reaction at 42 ℃, obtains placenta cDNA.Utilize the different primer of commentaries on classics (as shown in the table) of each gene, by 97 ℃ of 3 minutes, 1 circulations; 94 ℃ 30 seconds → 60 ℃ 30 seconds → 72 ℃ 1 minute, totally 35 circulations; 72 ℃ 10 minutes, pcr amplification is carried out in 1 circulation, obtains to contain the amplified production of each protein gene of complete open reading frame sequence.Amplified production is through sequence verification, and the sequence that records with embodiment 1 conforms to, and changes amplified production over to host cell with routine techniques subsequently, thereby obtains recombinant protein.(annotate:, can use the liver cDNA library of buying from GIBCO BRL company (catalog number (Cat.No.) :) 10422-012) as template for SP2114a.

Gene specific primer

Clone's title	Special primer 1 (5 ' → 3 ')	Special primer 2 (5 ' → 3 ')
			PP8153 PP8332 PP9177 PP9445 PP10199 PP10226 SP2114a	CGGAGGTTCTAGTGTCGGAG TTTCAGACCTGTTCCAAGGG CAGAGGAGCATCCCGTCTAC ACCAAATGAGGGACATGGAA GCAGTACTCCATGGTGCAGA AATGCACCCTGTTTTGAGAGA CCACTTCCACCAGAGACACA	GGACTGCTACCCATCCTGAA CTCTGCCTCCACTCACACTG GCTAGCCAGCTCTGTGGAGT CCAAGCCTGACTCTCTTTGC GTTTGCTCCCAGCTGTCTTC AGATCAACTTGAGGCCAGGA GCAAGGTTTCTTCAACTGGC

Embodiment 3:cDNA cloned sequence is analyzed

1.PP8153

A: nucleotide sequence (SEQ ID NO:1) length: 2349bp

1 GTGGAAGTAG AAGGCGGTGG CTGAGGCGGT TCCGGAGGTT CTAGTGTCGG AGTTGGGTGC

61 AGGCAGGTGC CATGGGCCCG CTTGAGGCAC ACTGAGGGGA CGCGGGGCTG GGCCATGGCC

121 GGCGCTCGGG CCGCCGCCGC CGCTGCCTCG GCGGGGTCCT CGGCCTCTTC AGGCAACCAG

181 CCGCCTCAGG AGCTGGGGCT TGGGGAGCTG CTGGAGGAGT TCTCCCGGAC TCAGTACCGG

241 GCCAAGGATG GCAGCGGGAC CGGCGGCTCT AAGGTTGAGC GCATTGAGAA GAGATGTCTG

301 GAGCTGTTTG GCCGAGACTA CTGTTTCAGC GTGATTCCAA ACACGAATGG GGATATCTGT

361 GGCCACTATC CCCGGCACAT CGTGTTCCTG GAGTATGAGA GTTCTGAGAA GGAGAAAGAC

421 ACGTTTGAGA GTACCGTACA GGTGAGCAAG TTGCAAGACC TCATCCACCG CAGCAAGATG

481 GCCCGGTGCA GAGGACGGTT TGTCTGCCCA GTAATCCTGT TCAAGGGCAA GCACATTTGC

541 AGGTCGGCCA CACTGGCTGG ATGGGGAGAG CTGTATGGAC GCTCAGGCTA CAACTATTTT

601 TTCTCAGGGG GTGCAGATGA TGCCTGGGCA GATGTGGAGG ACGTCACGGA GGAGGACTGT

661 GCTCTTCGAA GTGGTGACAC GCATCTTTTT GATAAGGTCA GAGGCTATGA CATCAAGCTG

721 CTTCGATACC TGTCAGTCAA ATACATCTGT GACCTGATGG TGGAGAACAA GAAGGTGAAG

781 TTTGGCATGA ATGTAACCTC CTCTGAGAAG GTGGACAAAG CCCAGCGCTA TGCCGACTTC

841 ACTCTCCTCT CCATCCCGTA TCCAGGCTGT GAATTTTTCA AGGAATATAA AGATCGGGAT

901 TACATGGCAG AAGGGCTCAT ATTTAACTGG AAGCAGGACT ACGTTGATGC CCCATTGAGC

961 ATCCCCGACT TCCTGACTCA CTCTCTGAAC ATTGACTGGA GCCAGTATCA GTGTTGGGAT

1021 CTGGTGCAAC AAACACAAAA CTACCTGAAG CTGCTGCTTT CCTTAGTTAA CAGTGATGAT

1081 GACAGCGGGC TGCTGGTACA CTGTATCTCA GGCTGGGATC GGACCCCCCT CTTCATCTCC

1141 CTCCTGCGCC TTTCCTTGTG GGCTGATGGG CTCATCCACA CGTCCCTGAA GCCCACTGAG

1201 ATCCTCTACC TCACTGTGGC CTATGACTGG TTCCTCTTCG GGCACATGTT GGTAGATCGG

1261 CTCAGCAAAG GGGAGGAGAT TTTCTTCTTC TGCTTCAATT TTTTGAAGCA TATTACCTCC

1321 GAGGAGTTCT CTGCTCTGAA GACCCAGAGG AGGAAGAGTT TGCCAGCCCG GGATGGAGGC

1381 TTCACCCTGG AAGACATCTG CATGCTGAGA CGAAAGGACC GTGGCAGCAC CACCAGCCTT

1441 GGCAGCGACT TCTCCCTGGT CATGGAGAGT TCCCCAGGAG CCACTGGGAG CTTCACCTAT

1501 GAAGGCCGTG GAGCTGGTCC CAGCAGGAGC GCCAACTCAG GCAGCTTGAA GGAAGAGCCA

1561 CTCATCCTCT CCACAGAGTG TCCTCTGGAA CCGGCCACAA CCCTCAGAGG ACCGCTTGCC

1621 TTCCCAGCAG GGGCTGGCGG AAGCCAGGTC TTCCAGCTCC TCTTCCTCAA ACCATTCTGA

1681 TAACTTTTTC AGGATGGGTA GCAGTCCCCT GGAGGTCCCC AAACCCAGGC TTGCAGCCCT

1741 GAGTGATCGA GAGACTCGGC TGCAGGAGGT GCGCTCAGCC TTCTTGGCTG CGTACAGCAG

1801 CACAGTGGGG CTTCGGGCAG TAGCCCCCAG TCCTTCCGGT GCCATCGGGG GCCTGCTGGA

1861 GCAATTTGCC CGTGGTGTTG GACTCCGGAG CATCAGCAGC AATGCCTTGT GAAGAAGCCA

1921 GCCCATGACA TTTTCCTGCT CCTCTCTCAG CTGAGCCCTT AGCAGAGAAT CAAAGCCATG

1981 CCTGGCCGAA GGGGTACTTC CAGGTCAGGG GAAATTTCAG TCCCCCATCT CCATCATGAA

2041 CATGGCAGCC CCAAAGCTGA GCAAGGCCAA AGACAGGGTT TTCCAACCCC CAGCCTCTTG

2101 ACTGGTGACC ACCACCCCTT CTTGTCACTG TCTCCCACCC ACCCCATCTT TGCTGGGATT

2161 CCCATCAACT CTCAGAACTG TGTGGGGTTT CCCTGGGGCC TTGTGGAAGC CATGACTTCA

2221 CAAAGACCCT ACCTGTCAGT TCTTGTTTCT GGGGAGGAGG GATCACCTGC ACTGAGAATG

2281 AGGCAGTTTG ACACAGATCA CAAAATAAAA TCAAAGTCTT TTTGAATAGC CAAAAAAAAA

2341 AAAAAAAAA

B: aminoacid sequence (SEQ ID NO:2) length: 521 amino acid

1 MAGARAAAAA ASAGSSASSG NQPPQELGLG ELLEEFSRTQ YRAKDGSGTG GSKVERIEKR

61 CLELFGRDYC FSVIPNTNGD ICGHYPRHIV FLEYESSEKE KDTFESTVQV SKLQDLIHRS

121 KMARCRGRFV CPVILFKGKH ICRSATLAGW GELYGRSGYN YFFSGGADDA WADVEDVTEE

181 DCALRSGDTH LFDKVRGYDI KLLRYLSVKY ICDLMVENKK VKFGMNVTSS EKVDKAQRYA

241 DFTLLSIPYP GCEFFKEYKD RDYMAEGLIF NWKQDYVDAP LSIPDFLTHS LNIDWSQYQC

301 WDLVQQTQNY LKLLLSLVNS DDDSGLLVHC ISGWDRTPLF ISLLRLSLWA DGLIHTSLKP

361 TEILYLTVAY DWFLFGHMLV DRLSKGEEIF FFCFNFLKHI TSEEFSALKT QRRKSLPARD

421 GGFTLEDICM LRRKDRGSTT SLGSDFSLVM ESSPGATGSF TYEGRGAGPS RSANSGSLKE

481 EPLILSTECP LEPATTLRGP LAFPAGAGGS QVFQLLFLKP F

C. Nucleotide and amino acid composite sequence (SEQ ID NO:3)

Clone number: PP8153

Start code: 115ATG stops coding: 1678TGA

Protein molecular weight: 58350.05

1 GTG GAA GTA GAA GGC GGT GGC TGA GGC GGT TCC GGA GGT TCT AGT GTC 48

49 GGA GTT GGG TGC AGG CAG GTG CCA TGG GCC CGC TTG AGG CAC ACT GAG 96

97 GGG ACG CGG GGC TGG GCC ATG GCC GGC GCT CGG GCC GCC GCC GCC GCT 144

1 Met Ala Gly Ala Arg Ala Ala Ala Ala Ala 10

145 GCC TCG GCG GGG TCC TCG GCC TCT TCA GGC AAC CAG CCG CCT CAG GAG 192

11 Ala Ser Ala Gly Ser Ser Ala Ser Ser Gly Asn Gln Pro Pro Gln Glu 26

193 CTG GGG CTT GGG GAG CTG CTG GAG GAG TTC TCC CGG ACT CAG TAC CGG 240

27 Leu Gly Leu Gly Glu Leu Leu Glu Glu Phe Ser Arg Thr Gln Tyr Arg 42

241 GCC AAG GAT GGC AGC GGG ACC GGC GGC TCT AAG GTT GAG CGC ATT GAG 288

43 Ala Lys Asp Gly Ser Gly Thr Gly Gly Ser Lys Val Glu Arg Ile Glu 58

289 AAG AGA TGT CTG GAG CTG TTT GGC CGA GAC TAC TGT TTC AGC GTG ATT 336

59 Lys Arg Cys Leu Glu Leu Phe Gly Arg Asp Tyr Cys Phe Ser Val Ile 74

337 CCA AAC ACG AAT GGG GAT ATC TGT GGC CAC TAT CCC CGG CAC ATC GTG 384

75 Pro Asn Thr Asn Gly Asp Ile Cys Gly His Tyr Pro Arg His Ile Val 90

385 TTC CTG GAG TAT GAG AGT TCT GAG AAG GAG AAA GAC ACG TTT GAG AGT 432

91 Phe Leu Glu Tyr Glu Ser Ser Glu Lys Glu Lys Asp Thr Phe Glu Ser 106

433 ACC GTA CAG GTG AGC AAG TTG CAA GAC CTC ATC CAC CGC AGC AAG ATG 480

107 Thr Val Gln Val Ser Lys Leu Gln Asp Leu Ile His Arg Ser Lys Met 122

481 GCC CGG TGC AGA GGA CGG TTT GTC TGC CCA GTA ATC CTG TTC AAG GGC 528

123 Ala Arg Cys Arg Gly Arg Phe Val Cys Pro Val Ile Leu Phe Lys Gly 138

529 AAG CAC ATT TGC AGG TCG GCC ACA CTG GCT GGA TGG GGA GAG CTG TAT 576

139 Lys His Ile Cys Arg Ser Ala Thr Leu Ala Gly Trp Gly Glu Leu Tyr 154

577 GGA CGC TCA GGC TAC AAC TAT TTT TTC TCA GGG GGT GCA GAT GAT GCC 624

155 Gly Arg Ser Gly Tyr Asn Tyr Phe Phe Ser Gly Gly Ala Asp Asp Ala 170

625 TGG GCA GAT GTG GAG GAC GTC ACG GAG GAG GAC TGT GCT CTT CGA AGT 672

171 Trp Ala Asp Val Glu Asp Val Thr Glu Glu Asp Cys Ala Leu Arg Ser 186

673 GGT GAC ACG CAT CTT TTT GAT AAG GTC AGA GGC TAT GAC ATC AAG CTG 720

187 Gly Asp Thr His Leu Phe Asp Lys Val Arg Gly Tyr Asp Ile Lys Leu 202

721 CTT CGA TAC CTG TCA GTC AAA TAC ATC TGT GAC CTG ATG GTG GAG AAC 768

203 Leu Arg Tyr Leu Ser Val Lys Tyr Ile Cys Asp Leu Met Val Glu Asn 218

769 AAG AAG GTG AAG TTT GGC ATG AAT GTA ACC TCC TCT GAG AAG GTG GAC 816

219 Lys Lys Val Lys Phe Gly Met Asn Val Thr Ser Ser Glu Lys Val Asp 234

817 AAA GCC CAG CGC TAT GCC GAC TTC ACT CTC CTC TCC ATC CCG TAT CCA 864

235 Lys Ala Gln Arg Tyr Ala Asp Phe Thr Leu Leu Ser Ile Pro Tyr Pro 250

865 GGC TGT GAA TTT TTC AAG GAA TAT AAA GAT CGG GAT TAC ATG GCA GAA 912

251 Gly Cys Glu Phe Phe Lys Glu Tyr Lys Asp Arg Asp Tyr Met Ala Glu 266

913 GGG CTC ATA TTT AAC TGG AAG CAG GAC TAC GTT GAT GCC CCA TTG AGC 960

267 Gly Leu Ile Phe Asn Trp Lys Gln Asp Tyr Val Asp Ala Pro Leu Ser 282

961 ATC CCC GAC TTC CTG ACT CAC TCT CTG AAC ATT GAC TGG AGC CAG TAT 1008

283 Ile Pro Asp Phe Leu Thr His Ser Leu Asn Ile Asp Trp Ser Gln Tyr 298

1009 CAG TGT TGG GAT CTG GTG CAA CAA ACA CAA AAC TAC CTG AAG CTG CTG 1056

299 Gln Cys Trp Asp Leu Val Gln Gln Thr Gln Asn Tyr Leu Lys Leu Leu 314

1057 CTT TCC TTA GTT AAC AGT GAT GAT GAC AGC GGG CTG CTG GTA CAC TGT 1104

315 Leu Ser Leu Val Asn Ser Asp Asp Asp Ser Gly Leu Leu Val His Cys 330

1105 ATC TCA GGC TGG GAT CGG ACC CCC CTC TTC ATC TCC CTC CTG CGC CTT 1152

331 Ile Ser Gly Trp Asp Arg Thr Pro Leu Phe Ile Ser Leu Leu Arg Leu 346

1153 TCC TTG TGG GCT GAT GGG CTC ATC CAC ACG TCC CTG AAG CCC ACT GAG 1200

347 Ser Leu Trp Ala Asp Gly Leu Ile His Thr Ser Leu Lys Pro Thr Glu 362

1201 ATC CTC TAC CTC ACT GTG GCC TAT GAC TGG TTC CTC TTC GGG CAC ATG 1248

363 Ile Leu Tyr Leu Thr Val Ala Tyr Asp Trp Phe Leu Phe Gly His Met 378

1249 TTG GTA GAT CGG CTC AGC AAA GGG GAG GAG ATT TTC TTC TTC TGC TTC 1296

379 Leu Val Asp Arg Leu Ser Lys Gly Glu Glu Ile Phe Phe Phe Cys Phe 394

1297 AAT TTT TTG AAG CAT ATT ACC TCC GAG GAG TTC TCT GCT CTG AAG ACC 1344

395 Asn Phe Leu Lys His Ile Thr Ser Glu Glu Phe Ser Ala Leu Lys Thr 410

1345 CAG AGG AGG AAG AGT TTG CCA GCC CGG GAT GGA GGC TTC ACC CTG GAA 1392

411 Gln Arg Arg Lys Ser Leu Pro Ala Arg Asp Gly Gly Phe Thr Leu Glu 426

1393 GAC ATC TGC ATG CTG AGA CGA AAG GAC CGT GGC AGC ACC ACC AGC CTT 1440

427 Asp Ile Cys Met Leu Arg Arg Lys Asp Arg Gly Ser Thr Thr Ser Leu 442

1441 GGC AGC GAC TTC TCC CTG GTC ATG GAG AGT TCC CCA GGA GCC ACT GGG 1488

443 Gly Ser Asp Phe Ser Leu Val Met Glu Ser Ser Pro Gly Ala Thr Gly 458

1489 AGC TTC ACC TAT GAA GGC CGT GGA GCT GGT CCC AGC AGG AGC GCC AAC 1536

459 Ser Phe Thr Tyr Glu Gly Arg Gly Ala Gly Pro Ser Arg Ser Ala Asn 474

1537 TCA GGC AGC TTG AAG GAA GAG CCA CTC ATC CTC TCC ACA GAG TGT CCT 1584

475 Ser Gly Ser Leu Lys Glu Glu Pro Leu Ile Leu Ser Thr Glu Cys Pro 490

1585 CTG GAA CCG GCC ACA ACC CTC AGA GGA CCG CTT GCC TTC CCA GCA GGG 1632

491 Leu Glu Pro Ala Thr Thr Leu Arg Gly Pro Leu Ala Phe Pro Ala Gly 506

1633 GCT GGC GGA AGC CAG GTC TTC CAG CTC CTC TTC CTC AAA CCA TTC TGA 1680

507 Ala Gly Gly Ser Gln Val Phe Gln Leu Leu Phe Leu Lys Pro Phe *** 522

1681 TAA CTT TTT CAG GAT GGG TAG CAG TCC CCT GGA GGT CCC CAA ACC CAG 1728

1729 GCT TGC AGC CCT GAG TGA TCG AGA GAC TCG GCT GCA GGA GGT GCG CTC 1776

1777 AGC CTT CTT GGC TGC GTA CAG CAG CAC AGT GGG GCT TCG GGC AGT AGC 1824

1825 CCC CAG TCC TTC CGG TGC CAT CGG GGG CCT GCT GGA GCA ATT TGC CCG 1872

1873 TGG TGT TGG ACT CCG GAG CAT CAG CAG CAA TGC CTT GTG AAG AAG CCA 1920

1921 GCC CAT GAC ATT TTC CTG CTC CTC TCT CAG CTG AGC CCT TAG CAG AGA 1968

1969 ATC AAA GCC ATG CCT GGC CGA AGG GGT ACT TCC AGG TCA GGG GAA ATT 2016

2017 TCA GTC CCC CAT CTC CAT CAT GAA CAT GGC AGC CCC AAA GCT GAG CAA 2064

2065 GGC CAA AGA CAG GGT TTT CCA ACC CCC AGC CTC TTG ACT GGT GAC CAC 2112

2113 CAC CCC TTC TTG TCA CTG TCT CCC ACC CAC CCC ATC TTT GCT GGG ATT 2160

2161 CCC ATC AAC TCT CAG AAC TGT GTG GGG TTT CCC TGG GGC CTT GTG GAA 2208

2209 GCC ATG ACT TCA CAA AGA CCC TAC CTG TCA GTT CTT GTT TCT GGG GAG 2256

2257 GAG GGA TCA CCT GCA CTG AGA ATG AGG CAG TTT GAC ACA GAT CAC AAA 2304

2305 ATA AAA TCA AAG TCT TTT TGA ATA GCC AAA AAA AAA AAA AAA AAA 2349

2.PP8332

A: nucleotide sequence (SEQ ID NO:4) length: 1771bp

1 GCCTGGGGCG TCCCCGCGAA GCCTGGGCCT GTCAGGCGGT TCCGTCCGGG TCTCGGCCAC

61 CGTCGAGTTC CGTCGAGTTC CGTCCCGGCC CTGCTCACAG CAGCGCCCTC GGAGCGCCCA

121 GCACCTGCGG CCGGCCAGGC AGCGCGATCC TGCGGCGTCT GGCCATCCCG AATGCTATGG

181 CCGCCGTCGC CGTCTTGCGG GCCTTCGGGG CAAGTGGGCC CATGTGTCTC CGGCGCGGCC

241 CCTGGGCCCA GCTCCCCGCC CGCTTCTGCA GCCGGGACCC GGCCGGGGCG GGGCGGCGGG

301 AGTCGGAGCC GCGGCCCACC AGCGCGCGGC AGCTGGACGG CATAAGGAAC ATCGTCTTGA

361 GCAATCCCAA GAAGAGGAAC ACGTTGTCAC TTGCAATGCT GAAATCTCTC CAAAGTGACA

421 TTCTTCATGA CGCTGACAGC AACGATCTGA AAGTCATTAT CATCTCGGCT GAGGGGCCTG

481 TGTTTTCTTC TGGGCATGAC TTAAAGGAGC TGACAGAGGA GCAAGGCCGT GATTACCATG

541 CCGAAGTATT TCAGACCTGT TCCAAGGGTC TCGCTCTGTC GCCCAGGCTG GATTACAGTG

601 GCATGATCTC GGCTCACTGC AACCTCTGCC TCCCGGGTTC AAGCAATTCT CCTGCCTCAG

661 CCTCCTGAGT AGCTGGGACT ACAGGTCATG ATGCACATCC GGAACCACCC CGTCCCCGTC

721 ATTGCCATGG TCAATGGCCT GGCCACGGCT GCCGGCTGTC AACTGGTTGC CAGCTGCGAC

781 ATTGCCGTGG CGAGCGACAA GTCCTCTTTT GCCACTCCTG GGGTGAACGT CGGGCTCTTC

841 TGTTCTACCC CTGGGGTTGC CTTGGCAAGA GCAGTGCCTA GAAAGGTGGC CTTGGAGATG

901 CTCTTTACTG GTGAGCCCAT TTCTGCCCAG GAGGCCCTGC TCCACGGGCT GCTTAGCAAG

961 GTGGTGCCAG AGGCGGAGCT GCAGGAGGAG ACCATGCGGA TCGCTAGGAA GATCGCATCG

1021 CTGAGCCGTC CGGTGGTGTC CCTGGGCAAA GCCACCTTCT ACAAGCAGCT GCCCCAGGAC

1081 CTGGGGACGG CTTACTACCT CACCTCCCAG GCCATGGTGG ACAACCTGGC CCTGCGGGAC

1141 GGGCAGGAGG GCATCACGGC CTTCCTCCAG AAGAGAAAAC CTGTCTGGTC ACACGAGCCA

1201 GTGTGAGTGG AGGCAGAGGA GTGAGGCCCA CGGGCAGCGC CCAGGAGCCC ACCTTCCCCT

1261 CTGGCCCAGC CACCACTGCC TCTCAGCTTC AACAGGTGAC AGGCTGCTTT CGTGACTTGA

1321 TATTGGTGTC ATAGCATTTG GCCTACATTA AAAGCCACAA TTTCATGGGG AAAGGACAAA

1381 ATGGAGAGTG ACTGAGGTGC TGACCTCAGT GCAAGGCTGG TGAACCCTGC AGCGGGCCAG

1441 CTATGGTGGG AAGCCTGGCA TTTGGGGTGC TCCTTGCAAC GTCTTAAGCA AGCGACCCCC

1501 CTGACATAGC AAAAGGTGGC AACCCATGGA GGCAGAAAGA AGGACGCCAG CCTGACCCTT

1561 ATCTTGAAAC GTCCTAAGCA GAGTTAATCC TGGCTGCTCA GGAGAGGCGA CACATTTCAA

1621 ATCTCCACGA GATATTCTCC ACACAGAAAA TCTTCTTGAT TCTATAGAGA CTTAATCATG

1681 CCTATGGCTT TGAATAATCT TATGTGATTT AAATAAATTA AATCTTTATA GAGACTGGAA

1741 AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA A

B: aminoacid sequence (SEQ ID NO:5) length: 172 amino acid

1 MMHIRNHPVP VIAMVNGLAT AAGCQLVASC DIAVASDKSS FATPGVNVGL FCSTPGVALA

61 RAVPRKVALE MLFTGEPISA QEALLHGLLS KVVPEAELQE ETMRIARKIA SLSRPVVSLG

121 KATFYKQLPQ DLGTAYYLTS QAMVDNLALR DGQEGITAFL QKRKPVWSHE PV

C. Nucleotide and amino acid composite sequence (SEQ ID NO:6)

Clone number: PP8332

Start code: 688ATG stops coding: 1204TGA

Protein molecular weight: 18409.48

1 GCC TGG GGC GTC CCC GCG AAG CCT GGG CCT GTC AGG CGG TTC CGT CCG 48

49 GGT CTC GGC CAC CGT CGA GTT CCG TCG AGT TCC GTC CCG GCC CTG CTC 96

97 ACA GCA GCG CCC TCG GAG CGC CGA GCA CCT GCG GCC GGC CAG GCA GCG 144

145 CGA TCC TGC GGC GTC TGG CCA TCC CGA ATG CTA TGG CCG CCG TCG CCG 192

193 TCT TGC GGG CCT TCG GGG CAA GTG GGC CCA TGT GTC TCC GGC GCG GCC 240

241 CCT GGG CCC AGC TCC CCG CCC GCT TCT GCA GCC GGG ACC CGG CCG GGG 288

289 CGG GGC GGC GGG AGT CGG AGC CGC GGC CCA CCA GCG CGC GGC AGC TGG 336

337 ACG GCA TAA GGA ACA TCG TCT TGA GCA ATC CCA AGA AGA GGA ACA CGT 384

385 TGT CAC TTG CAA TGC TGA AAT CTC TCC AAA GTG ACA TTC TTC ATG ACG 432

433 CTG ACA GCA ACG ATC TGA AAG TCA TTA TCA TCT CGG CTG AGG GGC CTG 480

481 TGT TTT CTT CTG GGC ATG ACT TAA AGG AGC TGA CAG AGG AGC AAG GCC 528

529 GTG ATT ACC ATG CCG AAG TAT TTC AGA CCT GTT CCA AGG GTC TCG CTC 576

577 TGT CGC CCA GGC TGG ATT ACA GTG GCA TGA TCT CGG CTC ACT GCA ACC 624

625 TCT GCC TCC CGG GTT CAA GCA ATT CTC CTG CCT CAG CCT CCT GAG TAG 672

673 CTG GGA CTA CAG GTC ATG ATG CAC ATC CGG AAC CAC CCC GTC CCC GTC 720

1 Met Met His Ile Arg Asn His Pro Val Pro Val 11

721 ATT GCC ATG GTC AAT GGC CTG GCC ACG GCT GCC GGC TGT CAA CTG GTT 768

12 Ile Ala Met Val Asn Gly Leu Ala Thr Ala Ala Gly Cys Gln Leu Val 27

769 GCC AGC TGC GAC ATT GCC GTG GCG AGC GAC AAG TCC TCT TTT GCC ACT 816

28 Ala Ser Cys Asp Ile Ala Val Ala Ser Asp Lys Ser Ser Phe Ala Thr 43

817 CCT GGG GTG AAC GTC GGG CTC TTC TGT TCT ACC CCT GGG GTT GCC TTG 864

44 Pro Gly Val Asn Val Gly Leu Phe Cys Ser Thr Pro Gly Val Ala Leu 59

865 GCA AGA GCA GTG CCT AGA AAG GTG GCC TTG GAG ATG CTC TTT ACT GGT 912

60 Ala Arg Ala Val Pro Arg Lys Val Ala Leu Glu Met Leu Phe Thr Gly 75

913 GAG CCC ATT TCT GCC CAG GAG GCC CTG CTC CAC GGG CTG CTT AGC AAG 960

76 Glu Pro Ile Ser Ala Gln Glu Ala Leu Leu His Gly Leu Leu Ser Lys 91

961 GTG GTG CCA GAG GCG GAG CTG CAG GAG GAG ACC ATG CGG ATC GCT AGG 1008

92 Val Val Pro Glu Ala Glu Leu Gln Glu Glu Thr Met Arg Ile Ala Arg 107

1009 AAG ATC GCA TCG CTG AGC CGT CCG GTG GTG TCC CTG GGC AAA GCC ACC 1056

108 Lys Ile Ala Ser Leu Ser Arg Pro Val Val Ser Leu Gly Lys Ala Thr 123

1057 TTC TAC AAG CAG CTG CCC CAG GAC CTG GGG ACG GCT TAC TAC CTC ACC 1104

124 Phe Tyr Lys Gln Leu Pro Gln Asp Leu Gly Thr Ala Tyr Tyr Leu Thr 139

1105 TCC CAG GCC ATG GTG GAC AAC CTG GCC CTG CGG GAC GGG CAG GAG GGC 1152

140 Ser Gln Ala Met Val Asp Asn Leu Ala Leu Arg Asp Gly Gln Glu Gly 155

1153 ATC ACG GCC TTC CTC CAG AAG AGA AAA CCT GTC TGG TCA CAC GAG CCA 1200

156 Ile Thr Ala Phe Leu Gln Lys Arg Lys Pro Val Trp Ser His Glu Pro 171

1201 GTG TGA GTG GAG GCA GAG GAG TGA GGC CCA CGG GCA GCG CCC AGG AGC 1248

172 Val *** 173

1249 CCA CCT TCC CCT CTG GCC CAG CCA CCA CTG CCT CTC AGC TTC AAC AGG 1296

1297 TGA CAG GCT GCT TTC GTG ACT TGA TAT TGG TGT CAT AGC ATT TGG CCT 1344

1345 ACA TTA AAA GCC ACA ATT TCA TGG GGA AAG GAC AAA ATG GAG AGT GAC 1392

1393 TGA GGT GCT GAC CTC AGT GCA AGG CTG GTG AAC CCT GCA GCG GGC CAG 1440

1441 CTA TGG TGG GAA GCC TGG CAT TTG GGG TGC TCC TTG CAA CGT CTT AAG 1488

1489 CAA GCG ACC CCC CTG ACA TAG CAA AAG GTG GCA ACC CAT GGA GGC AGA 1536

1537 AAG AAG GAC GCC AGC CTG ACC CTT ATC TTG AAA CGT CCT AAG CAG AGT 1584

1585 TAA TCC TGG CTG CTC AGG AGA GGC GAC ACA TTT CAA ATC TCC ACG AGA 1632

1633 TAT TCT CCA CAC AGA AAA TCT TCT TGA TTC TAT AGA GAC TTA ATC ATG 1680

1681 CCT ATG GCT TTG AAT AAT CTT ATG TGA TTT AAA TAA ATT AAA TCT TTA 1728

1729 TAG AGA CTG GAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA A 1771

3.PP9177

A: nucleotide sequence (SEQ ID NO:7) length: 2160bp

1 GCGTCTGCCA GCCGGCTTGG CTAGCGCGCG GCGGCCGTGG CTAAGGCTGC TACGAAGCGA

61 GCTTGGGAGG AGCAGCGGCC TGCGGGGCAG AGGAGCATCC CGTCTACCAG GTCCCAAGCG

121 GCCGTGGCCC GCGGGTCATG GCCAAAGGAG AAGGCGCCGA GAGCGGCTCC GCGGCGGGGC

181 TGCTACCCAC CAGCATCCTC CAAAGCACTG AACGCCCGGC CCAGGTGAAG AAAGAACCGA

241 AAAAGAAGAA ACAACAGTTG TCTGTTTGCA ACAAGCTTTG CTATGCACTT GGGGGAGCCC

301 CCTACCAGGT GACGGGCTGT GCCCTGGGTT TCTTCCTTCA GATCTACCTA TTGGATGTGG

361 CTCAGGTGGG CCCTTTCTCT GCCTCCATCA TCCTGTTTGT GGGCCGAGCC TGGGATGCCA

421 TCACAGACCC CCTGGTGGGC CTCTGCATCA GCAAATCCCC CTGGACCTGC CTGGGTCGCC

481 TTATGCCCTG GATCATCTTC TCCACGCCCC TGGCCGTCAT TGCCTACTTC CTCATCTGGT

541 TCGTGCCCGA CTTCCCACAC GGCCAGACCT ATTGGTACCT GCTTTTCTAT TGCCTCTTTG

601 AAACAATGGT CACGTGTTTC CATGTTCCCT ACTCGGCTCT CACCATGTTC ATCAGCACCG

661 AGCAGACTGA GCGGGATTCT GCCACCGCCT ATCGGATGAC TGTGGAAGTG CTGGGCACAG

721 TGCTGGGCAC GGCGATCCAG GGACAAATCG TGGGCCAAGC AGACACGCCT TGTTTCCAGG

781 ACCTCAATAG CTCTACAGTA GCTTCACAAA GTGCCAACCA TACACATGGC ACCACCTCAC

841 ACAGGGAAAC GCAAAAGGCA TACCTGCTGG CAGCGGGGGT CATTGTCTGT ATCTATATAA

901 TCTGTGCTGT CATCCTGATC CTGGGCGTGC GGGAGCAGAG AGAACCCTAT GAAGCCCAGC

961 AGTCTGAGCC AATCGCCTAC TTCCGGGGCC TACGGCTGGT CATGAGCCAC GGCCCATACA

1021 TCAAACTTAT TACTGGCTTC CTCTTCACCT CCTTGGCTTT CATGCTGGTG GAGGGGAACT

1081 TTGTCTTGTT TTGCACCTAC ACCTTGGGCT TCCGCAATGA ATTCCAGAAT CTACTCCTGG

1141 CCATCATGCT CTCGGCCACT TTAACCATTC CCATCTGGCA GTGGTTCTTG ACCCGGTTTG

1201 GCAAGAAGAC AGCTGTATAT GTTGGGATCT CATCAGCAGT GCCATTTCTC ATCTTGGTGG

1261 CCCTCATGGA GAGTAACCTC ATCATTACAT ATGCGGTAGC TGTGGCAGCT GGCATCAGTG

1321 TGGCAGCTGC CTTCTTACTA CCCTGGTCCA TGCTGCCTGA TGTCATTGAC GACTTCCATC

1381 TGAAGCAGCC CCACTTCCAT GGAACCGAGC CCATCTTCTT CTCCTTCTAT GTCTTCTTCA

1441 CCAAGTTTGC CTCTGGAGTG TCACTGGGCA TTTCTACCCT CAGTCTGGAC TTTGCAGGGT

1501 ACCAGACCCG TGGCTGCTCG CAGCCGGAAC GTGTCAAGTT TACACTGAAC ATGCTCGTGA

1561 CCATGGCTCC CATAGTTCTC ATCCTGCTGG GCCTGCTGCT CTTCAAAATG TACCCCATTG

1621 ATGAGGAGAG GCGGCGGCAG AATAAGAAGG CCCTGCAGGC ACTGAGGGAC GAGGCCAGCA

1681 GCTCTGGCTG CTCAGAAACA GACTCCACAG AGCTGGCTAG CATCCTCTAG GGCCCGCCAC

1741 GTTGCCCGAA GCCACCATGC AGAAGGCCAC AGAAGGGATC AGGACCTGTC TGCCGGCTTG

1801 CTGAGCAGCT GGACTGCAGG TGCTAGGAAG GGAACTGAAG ACTCAAGGAG GTGGCCCAGG

1861 ACACTTGCTG TGCTCACTGT GGGGCCGGCT GCTCTGTGGC CTCCTGCCTC CCCTCTGCCT

1921 GCCTGTGGGG CCAAGCCCTG GGGCTGCCAC TGTGAATATG CCAAGGACTG ATCGGGCCTA

1981 GCCCGGAACA CTAATGTAGA AACCTTTTTT TTACAGAGCC TAATTAATAA CTTAATGACT

2041 GTGTACATAG CAATGTGTGT GTATGTATAT GTCTGTGAGC TATTAATGTT ATTAATTTTC

2101 ATAAAAGCTG GAAAGCAAAA AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA

B: aminoacid sequence (SEQ ID NO:8) length: 530 amino acid

1 MAKGEGAESG SAAGLLPTSI LQSTERPAQV KKEPKKKKQQ LSVCNKLCYA LGGAPYQVTG

61 CALGFFLQIY LLDVAQVGPF SASIILFVGR AWDAITDPLV GLCISKSPWT CLGRLMPWII

121 FSTPLAVIAY FLIWFVPDFP HGQTYWYLLF YCLFETMVTC FHVPYSALTM FISTEQTERD

181 SATAYRMTVE VLGTVLGTAI QGQIVGQADT PCFQDLNSST VASQSANHTH GTTSHRETQK

241 AYLLAAGVIV CIYIICAVIL ILGVREQREP YEAQQSEPIA YFRGLRLVMS HGPYIKLITG

301 FLFTSLAFML VEGNFVLFCT YTLGFRNEFQ NLLLAIMLSA TLTIPIWQWF LTRFGKKTAV

361 YVGISSAVPF LILVALMESN LIITYAVAVA AGISVAAAFL LPWSMLPDVI DDFHLKQPHF

421 HGTEPIFFSF YVFFTKFASG VSLGISTLSL DFAGYQTRGC SQPERVKFTL NMLVTMAPIV

481 LILLGLLLFK MYPIDEERRR QNKKALQALR DEASSSGCSE TDSTELASIL

C. Nucleotide and amino acid composite sequence (SEQ ID NO:9)

Clone number: PP9177

Start code: 138ATG stops coding: 1728TAG

Protein molecular weight: 58620.36

1 GC GTC TGC CAG CCG GCT TGG CTA GCG CGC GGC GGC CGT GGC TAA GGC 47

48 TGC TAC GAA GCG AGC TTG GGA GGA GCA GCG GCC TGC GGG GCA GAG GAG 95

96 CAT CCC GTC TAC CAG GTC CCA AGC GGC CGT GGC CCG CGG GTC ATG GCC 143

1 Met Ala 2

144 AAA GGA GAA GGC GCC GAG AGC GGC TCC GCG GCG GGG CTG GTA CCC ACC 191

3 Lys Gly Glu Gly Ala Glu Ser Gly Ser Ala Ala Gly Leu Leu Pro Thr 18

192 AGC ATC CTC CAA AGC ACT GAA CGC CCG GCC CAG GTG AAG AAA GAA CCG 239

19 Ser Ile Leu Gln Ser Thr Glu Arg Pro Ala Gln Val Lys Lys Glu Pro 34

240 AAA AAG AAG AAA CAA CAG TTG TCT GTT TGC AAC AAG CTT TGC TAT GCA 287

35 Lys Lys Lys Lys Gln Gln Leu Ser Val Cys Asn Lys Leu Cys Tyr Ala 50

288 CTT GGG GGA GCC CCC TAC CAG GTG ACG GGC TGT GCC CTG GGT TTC TTC 335

51 Leu Gly Gly Ala Pro Tyr Gln Val Thr Gly Cys Ala Leu Gly Phe Phe 66

336 CTT CAG ATC TAC CTA TTG GAT GTG GCT CAG GTG GGC CCT TTC TCT GCC 383

67 Leu Gln Ile Tyr Leu Leu Asp Val Ala Gln Val Gly Pro Phe Ser Ala 82

384 TCC ATC ATC CTG TTT GTG GGC CGA GCC TGG GAT GCC ATC ACA GAC CCC 431

83 Ser Ile Ile Leu Phe Val Gly Arg Ala Trp Asp Ala Ile Thr Asp Pro 98

432 CTG GTG GGC CTC TGC ATC AGC AAA TCC CCC TGG ACC TGC CTG GGT CGC 479

99 Leu Val Gly Leu Cys Ile Ser Lys Ser Pro Trp Thr Cys Leu Gly Arg 114

480 CTT ATG CCC TGG ATC ATC TTC TCC ACG CCC CTG GCC GTC ATT GCC TAC 527

115 Leu Met Pro Trp Ile Ile Phe Ser Thr Pro Leu Ala Val Ile Ala Tyr 130

528 TTC CTC ATC TGG TTC GTG CCC GAC TTC CCA CAC GGC CAG ACC TAT TGG 575

131 Phe Leu Ile Trp Phe Val Pro Asp Phe Pro His Gly Gln Thr Tyr Trp 146

576 TAC CTG CTT TTC TAT TGC CTC TTT GAA ACA ATG GTC ACG TGT TTC CAT 623

147 Tyr Leu Leu Phe Tyr Cys Leu Phe Glu Thr Met Val Thr Cys Phe His 162

624 GTT CCC TAC TCG GCT CTC ACC ATG TTC ATC AGC ACC GAG CAG ACT GAG 671

163 Val Pro Tyr Ser Ala Leu Thr Met Phe Ile Ser Thr Glu Gln Thr Glu 178

672 CGG GAT TCT GCC ACC GCC TAT CGG ATG ACT GTG GAA GTG CTG GGC ACA 719

179 Arg Asp Ser Ala Thr Ala Tyr Arg Met Thr Val Glu Val Leu Gly Thr 194

720 GTG CTG GGC ACG GCG ATC CAG GGA CAA ATC GTG GGC CAA GCA GAC ACG 767

195 Val Leu Gly Thr Ala Ile Gln Gly Gln Ile Val Gly Gln Ala Asp Thr 210

768 CCT TGT TTC CAG GAC CTC AAT AGC TCT ACA GTA GCT TCA CAA AGT GCC 815

211 Pro Cys Phe Gln Asp Leu Asn Ser Ser Thr Val Ala Ser Gln Ser Ala 226

816 AAC CAT ACA CAT GGC ACC ACC TCA CAC AGG GAA ACG CAA AAG GCA TAC 863

227 Asn His Thr His Gly Thr Thr Ser His Arg Glu Thr Gln Lys Ala Tyr 242

864 CTG CTG GCA GCG GGG GTC ATT GTC TGT ATC TAT ATA ATC TGT GCT GTC 911

243 Leu Leu Ala Ala Gly Val Ile Val Cys Ile Tyr Ile Ile Cys Ala Val 258

912 ATC CTG ATC CTG GGC GTG CGG GAG CAG AGA GAA CCC TAT GAA GCC CAG 959

259 Ile Leu Ile Leu Gly Val Arg Glu Gln Arg Glu Pro Tyr Glu Ala Gln 274

960 CAG TCT GAG CCA ATC GCC TAC TTC CGG GGC CTA CGG CTG GTC ATG AGC 1007

275 Gln Ser Glu Pro Ile Ala Tyr Phe Arg Gly Leu Arg Leu Val Met Ser 290

1008 CAC GGC CCA TAC ATC AAA CTT ATT ACT GGC TTC CTC TTC ACC TCC TTG 1055

291 His Gly Pro Tyr Ile Lys Leu Ile Thr Gly Phe Leu Phe Thr Ser Leu 306

1056 GCT TTC ATG CTG GTG GAG GGG AAC TTT GTC TTG TTT TGC ACC TAC ACC 1103

307 Ala Phe Met Leu Val Glu Gly Asn Phe Val Leu Phe Cys Thr Tyr Thr 322

1104 TTG GGC TTC CGC AAT GAA TTC CAG AAT CTA CTC CTG GCC ATC ATG CTC 1151

323 Leu Gly Phe Arg Asn Glu Phe Gln Asn Leu Leu Leu Ala Ile Met Leu 338

1152 TCG GCC ACT TTA ACC ATT CCC ATC TGG CAG TGG TTC TTG ACC CGG TTT 1199

339 Ser Ala Thr Leu Thr Ile Pro Ile Trp Gln Trp Phe Leu Thr Arg Phe 354

1200 GGC AAG AAG ACA GCT GTA TAT GTT GGG ATC TCA TCA GCA GTG CCA TTT 1247

355 Gly Lys Lys Thr Ala Val Tyr Val Gly Ile Ser Ser Ala Val Pro Phe 370

1248 CTC ATC TTG GTG GCC CTC ATG GAG AGT AAC CTC ATC ATT ACA TAT GCG 1295

371 Leu Ile Leu Val Ala Leu Met Glu Ser Asn Leu Ile Ile Thr Tyr Ala 386

1296 GTA GCT GTG GCA GCT GGC ATC AGT GTG GCA GCT GCC TTC TTA CTA CCC 1343

387 Val Ala Val Ala Ala Gly Ile Ser Val Ala Ala Ala Phe Leu Leu Pro 402

1344 TGG TCC ATG CTG CCT GAT GTC ATT GAC GAC TTC CAT CTG AAG CAG CCC 1391

403 Trp Ser Met Leu Pro Asp Val Ile Asp Asp Phe His Leu Lys Gln Pro 418

1392 CAC TTC CAT GGA ACC GAG CCC ATC TTC TTC TCC TTC TAT GTC TTC TTC 1439

419 His Phe His Gly Thr Glu Pro Ile Phe Phe Ser Phe Tyr Val Phe Phe 434

1440 ACC AAG TTT GCC TCT GGA GTG TCA CTG GGC ATT TCT ACC CTC AGT CTG 1487

435 Thr Lys Phe Ala Ser Gly Val Ser Leu Gly Ile Ser Thr Leu Ser Leu 450

1488 GAC TTT GCA GGG TAC CAG ACC CGT GGC TGC TCG CAG CCG GAA CGT GTC 1535

451 Asp Phe Ala Gly Tyr Gln Thr Arg Gly Cys Ser Gln Pro Glu Arg Val 466

1536 AAG TTT ACA CTG AAC ATG CTC GTG ACC ATG GCT CCC ATA GTT CTC ATC 1583

467 Lys Phe Thr Leu Asn Met Leu Val Thr Met Ala Pro Ile Val Leu Ile 482

1584 CTG CTG GGC CTG CTG CTC TTC AAA ATG TAC CCC ATT GAT GAG GAG AGG 1631

483 Leu Leu Gly Leu Leu Leu Phe Lys Met Tyr Pro Ile Asp Glu Glu Arg 498

1632 CGG CGG CAG AAT AAG AAG GCC CTG CAG GCA CTG AGG GAC GAG GCC AGC 1679

499 Arg Arg Gln Asn Lys Lys Ala Leu Gln Ala Leu Arg Asp Glu Ala Ser 514

1680 AGC TCT GGC TGC TCA GAA AGA GAG TCG AGA GAG GTG GCT AGC ATC CTC 1727

515 Ser Ser Gly Cys Ser Glu Thr Asp Ser Thr Glu Leu Ala Ser Ile Leu 530

1728 TAG GGC CCG CCA CGT TGC CCG AAG CCA CCA TGC AGA AGG CCA CAG AAG 1775

531 *** 531

1776 GGA TCA GGA CCT GTC TGC CGG CTT GCT GAG CAG CTG GAC TGC AGG TGC 1823

1824 TAG GAA GGG AAC TGA AGA CTC AAG GAG GTG GCC CAG GAC ACT TGC TGT 1871

1872 GCT CAC TGT GGG GCC GGC TGC TCT GTG GCC TCC TGC CTC CCC TCT GCC 1919

1920 TGC CTG TGG GGC CAA GCC CTG GGG CTG CCA CTG TGA ATA TGC CAA GGA 1967

1968 CTG ATC GGG CCT AGC CCG GAA CAC TAA TGT AGA AAC CTT TTT TTT ACA 2015

2016 GAG CCT AAT TAA TAA CTT AAT GAC TGT GTA CAT AGC AAT GTG TGT GTA 2063

2064 TGT ATA TGT CTG TGA GCT ATT AAT GTT ATT AAT TTT CAT AAA AGC TGG 2111

2112 AAA GCA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA AAA 2159

2160 A 2160

4.PP9445

A: nucleotide sequence (SEQ ID NO:10) length: 1831bp

1 GCCGCCGCGG AGCGAGGTTG ACTGGAGAGA GCGCCTGGGC GCAGAAGGGT TAACGGGCCA

61 CCGGGGGCTC GCAGAGCAGG AGGGTGCTCT CGGACGGTGT GTCCCCCACT GCACTCCTGA

121 ACTTGGAGGA CAGGGTCGCC GCGAGGGACG CAGGTGGGTG CCCTTGATCC AGCTCAGCCC

181 GATGGCAGAA GAGGTTGACA AAAAAGAAAG ACACCTGTTG GGGTGGCCTG CCAGACCCAG

241 GAGTGGAGGG CTCTGTGAGG GCCCGGGAAT TCGGACTCAG GACAGGGATT CTCCATGGCT

301 AGGCCCAGAA ACACAGGGTC CAACCACTCT CCAGCAGGGA GACCTGGGGG TGAAGGGGTG

361 AGCCCTGCGC AGGTCTCTGT TCCTTGGTCT TCACTGGGCA GTGTGGAGAG GTGTGGCCAG

421 GAGGAGCCCG CGTTTGTCCA GACCAGGGTC TACTCTGGCA CCAGAGTGAC CACCTCTGAC

481 CTCTCCTTTC CTCGTCCTGG GCCGGGAACG ACACCAAATG AGGGACATGG AAAGGGCTGG

541 AGTAACAAGA GTCAGGCAGA GCCTGAAGAC TTGGGTGGAA CATGGGCCCT TCTCTGGAGA

601 TCCTGGCCTC CCCCGTTCAG TCAGGGTGGA GTTGCTGACC TTAGTGGCCG GCCCAGCCAG

661 GGGAAGGAGT GGCCATCGGC AACCCCCACC CCAACCCCAA TCCCTGAGGC GCCCGCTCTG

721 GCTCAGCCAC TCTGACCCCT CCCTCAAATT CCGAACCCTA GGTCTCAGGG AGGGCAGTGG

781 GGCTGAGTGT CTGCCCCCAG GCACATTCCT ACCCTTCTCT TGGTCATTTT CTGCCCCAGA

841 GCTGGCCCAC CTCAGCAATG CGAGGGCTCC CTGGATTCCT CTCCCGGGTG CCTTTCAGAT

901 CCAACAGAAA CAGATTTTTT TTTTCCTGGA AAGCAGAACT AAGAGTGGGA TGAGGAGCAG

961 GGGTGGGAAG GACTCAAAGT GAGAAGAAGG GGGCAAAGAG AGTCAGGCTT GGTGGCTGGG

1021 GTGGCTTCCA AGCCTCACTT CTCCAGTGTT CAAAGCTGAA CTTCAGATGG ACTTCCCGGC

1081 TCTTCAGAAT GAGAGGCCTG TGGCTGGGGC ATGAGGCAGC CCCGGCTGCA CCTCTCCTTC

1141 CCGCTTCCCC AGCTGGTAGA GACGCACAGG AAACAAGCCC TCACTGAACC AACTCCAGAT

1201 GCTGGCACCC AGAGTGGGTG TTACATTGCC GGCTTCTTCT CTAGAGATTA AACCGTCAAC

1261 CCATTTAGCT TATCCCTTGG CCAAAAAGTG TATGAGATGT GCCTGGATGT TCCCTAAAGA

1321 GCTTATCTAA GAAGGGAAGA GAAAGCCGGG AGGCAAGTAG GACAGAGAGA TGACTGGGGA

1381 AGGTCTTGTG TCTGGAAGAC CCAAGGAAGG GGCTTCTGGT GGGTCCTCAG AGAGAGTGTC

1441 TGGCGCATCC TCAGTGGAGC CTTCCTCCTC TACTTTCTAG GCACCTCTGG GAGGGCAGGA

1501 GTGGGAGCAG ATGACAACCA TTTTAGAAGG AGCCCTCTGG CTGGGTGCGG TGGCTCACAC

1561 CTGTCATCCC AGCACTTTGG GAGGCCAAGG CAGGAGAAGC GCTTGAGGCC TGGAGTTCAA

1621 GACCAGCCTG TGCAATTTAG CTGGATCCCA TCTCCACCAA AAAATACCAA AATTAGCTGG

1681 GTGTGGTGGT GCACGCATGT AGTCCCACCT ACTCAGGAGG CTGAGGAAGG AGAGCCTGTG

1741 AGTTTGAGGC TGCAATGAGC TTTGGTGGCA CCACTGCCCT CCAGCCTGGA TGACAGAGTG

1801 AGATCTCCAT CTCAAAAAAA AAAAAAAAAA A

B: aminoacid sequence (SEQ ID NO:11) length: 154 amino acid

1 MRDMERAGVT RVRQSLKTWV EHGPFSGDPG LPRSVRVELL TLVAGPARGR SGHRQPPPQP

61 QSLRRPLWLS HSDPSLKFRT LGLREGSGAE CLPPGTFLPF SWSFSAPELA HLSNARAPWI 121 PLPGAFQIQQ KQIFFFLESR TKSGMRSRGG KDSK

C. Nucleotide and amino acid composite sequence (SEQ ID NO:12)

Clone number: PP9445

Start code: 518ATG stops coding: 980TGA

Protein molecular weight: 17150.75

1 G CCG CCG CGG AGC GAG GTT GAC TGG AGA GAG CGC CTG GGC GCA GAA 46

47 GGG TTA ACG GGC CAC CGG GGG CTC GCA GAG CAG GAG GGT GCT CTC GGA 94

95 CGG TGT GTC CCC CAC TGC ACT CCT GAA CTT GGA GGA CAG GGT CGC CGC 142

143 GAG GGA CGC AGG TGG GTG CCC TTG ATC CAG CTC AGC CCG ATG GCA GAA 190

191 GAG GTT GAC AAA AAA GAA AGA CAC CTG TTG GGG TGG CCT GCC AGA CCC 238

239 AGG AGT GGA GGG CTC TGT GAG GGC CCG GGA ATT CGG ACT CAG GAC AGG 286

287 GAT TCT CCA TGG CTA GGC CCA GAA ACA CAG GGT CCA ACC ACT CTC CAG 334

335 CAG GGA GAC CTG GGG GTG AAG GGG TGA GCC CTG CGC AGG TCT CTG TTC 382

383 CTT GGT CTT CAC TGG GCA GTG TGG AGA GGT GTG GCC AGG AGG AGC CCG 430

431 CGT TTG TCC AGA CCA GGG TCT ACT CTG GCA CCA GAG TGA CCA CCT CTG 478

479 ACC TCT CCT TTC CTC GTC CTG GGC CGG GAA CGA CAC CAA ATG AGG GAC 526

1 Met Arg Asp 3

527 ATG GAA AGG GCT GGA GTA ACA AGA GTC AGG CAG AGC CTG AAG ACT TGG 574

4 Met Glu Arg Ala Gly Val Thr Arg Val Arg Gln Ser Leu Lys Thr Trp 19

575 GTG GAA CAT GGG CCC TTC TCT GGA GAT CCT GGC CTC CCC CGT TCA GTC 622

20 Val Glu His Gly Pro Phe Ser Gly Asp Pro Gly Leu Pro Arg Ser Val 35

623 AGG GTG GAG TTG CTG ACC TTA GTG GCC GGC CCA GCC AGG GGA AGG AGT 670

36 Arg Val Glu Leu Leu Thr Leu Val Ala Gly Pro Ala Arg Gly Arg Ser 51

671 GGC CAT CGG CAA CCC CCA CCC CAA CCC CAA TCC CTG AGG CGC CCG CTC 718

52 Gly His Arg Gln Pro Pro Pro Gln Pro Gln Ser Leu Arg Arg Pro Leu 67

719 TGG CTC AGC CAC TCT GAC CCC TCC CTC AAA TTC CGA ACC CTA GGT CTC 766

68 Trp Leu Ser His Ser Asp Pro Ser Leu Lys Phe Arg Thr Leu Gly Leu 83

767 AGG GAG GGC AGT GGG GCT GAG TGT CTG CCC CCA GGC ACA TTC CTA CCC 814

84 Arg Glu Gly Ser Gly Ala Glu Cys Leu Pro Pro Gly Thr Phe Leu Pro 99

815 TTC TCT TGG TCA TTT TCT GCC CCA GAG CTG GCC CAC CTC AGC AAT GCG 862

100 Phe Ser Trp Ser Phe Ser Ala Pro Glu Leu Ala His Leu Ser Asn Ala 115

863 AGG GCT CCC TGG ATT CCT CTC CCG GGT GCC TTT CAG ATC CAA CAG AAA 910

116 Arg Ala Pro Trp Ile Pro Leu Pro Gly Ala Phe Gln Ile Gln Gln Lys 131

911 CAG ATT TTT TTT TTC CTG GAA AGC AGA ACT AAG AGT GGG ATG AGG AGC 958

132 Gln Ile Phe Phe Phe Leu Glu Ser Arg Thr Lys Ser Gly Met Arg Ser 147

959 AGG GGT GGG AAG GAC TCA AAG TGA GAA GAA GGG GGC AAA GAG AGT CAG 1006

148 Arg Gly Gly Lys Asp Ser Lys *** 155

1007 GCT TGG TGG CTG GGG TGG CTT CCA AGC CTC ACT TCT CCA GTG TTC AAA 1054

1055 GCT GAA CTT CAG ATG GAC TTC CCG GCT CTT CAG AAT GAG AGG CCT GTG 1102

1103 GCT GGG GCA TGA GGC AGC CCC GGC TGC ACC TCT CCT TCC CGC TTC CCC 1150

1151 AGC TGG TAG AGA CGC ACA GGA AAC AAG CCC TCA CTG AAC CAA CTC CAG 1198

1199 ATG CTG GCA CCC AGA GTG GGT GTT ACA TTG CCG GCT TCT TCT CTA GAG 1246

1247 ATT AAA CCG TCA ACC CAT TTA GCT TAT CCC TTG GCC AAA AAG TGT ATG 1294

1295 AGA TGT GCC TGG ATG TTC CCT AAA GAG CTT ATC TAA GAA GGG AAG AGA 1342

1343 AAG CCG GGA GGC AAG TAG GAC AGA GAG ATG ACT GGG GAA GGT CTT GTG 1390

1391 TCT GGA AGA CCC AAG GAA GGG GCT TCT GGT GGG TCC TCA GAG AGA GTG 1438

1439 TCT GGC GCA TCC TCA GTG GAG CCT TCC TCC TCT ACT TTC TAG GCA CCT 1486

1487 CTG GGA GGG CAG GAG TGG GAG CAG ATG ACA ACC ATT TTA GAA GGA GCC 1534

1535 CTC TGG CTG GGT GCG GTG GCT CAC ACC TGT CAT CCC AGC ACT TTG GGA 1582

1583 GGC CAA GGC AGG AGA AGC GCT TGA GGC CTG GAG TTC AAG ACC AGC CTG 1630

1631 TGC AAT TTA GCT GGA TCC CAT CTC CAC CAA AAA ATA CCA AAA TTA GCT 1678

1679 GGG TGT GGT GGT GCA CGC ATG TAG TCC CAC CTA CTC AGG AGG CTG AGG 1726

1727 AAG GAG AGC CTG TGA GTT TGA GGC TGC AAT GAG CTT TGG TGG CAC CAC 1774

1775 TGC CCT CCA GCC TGG ATG ACA GAG TGA GAT CTC CAT CTC AAA AAA AAA 1822

1823 AAA AAA AAA 1831

5.PP10199

A: nucleotide sequence (SEQ ID NO:13) length: 1739bp

1 GTTCCAGAGC CACTTTTAAG ATTCTTCAAT TCCAAATGCA TGTCTTTTTT TAAAAAAAAG

61 AAAGAAAGAA AAATAAGTTT CTAATATTAG AGAAGTACAG CCCTGAATTG GGTTTTGTGT

121 CCACTGCTGG ACCCCATGAG GGCCAGGTGG AGTGGACCTC TGCAGCCCCA GTTGTGTGCA

181 CTCTCTGTTT GGTGCAAATT CCAGTTTGCT GGTTCTCAAT AGCAAGACCA GCCTGAGACC

241 ACCTGTCCTG CTCTTCCCAT GAGAGGGCCG AATGCTCCCA GCCTCCATGC CATGTCCTGT

301 TCCTGGGGTC CTGGGGGTCA TTGCAGCCTG TATGTGCTTC CTCCAGCCAG GGTGATCATC

361 GGGTGCCCCA GTGAGCCCCA GCACTGAGGG TCAGCCCCAG GCACTGTCAA AGGTGAGAGC

421 TCAGAGGCTG TGCCCAGAAA GAGAGGTGGG CCCTGCCTGC CCTGGACGGA GGGAGAGAGG

481 CTTCTCAGAG CCCGAGGCAT GAACCCTCAG GTGGGTCGTG GCCATAGTCA GATGATGGCT

541 GCTGGTGAGC TCAGTGACCA GGCGTCTTCA GGCAGCTCAT AAGTTTGAGA GGACACAGCC

601 TAAGGGAGGT TTGCTGGGGA GTAGCCCCAC TTCCACCCTG AATAGACAAG AGATGGTAAA

661 GCAGGTACCC AGCACTTAGT GCTTTCTTGG GGATATCGCG TGGGTCCCCG GGGGCCTGGG

721 TGCCCGAAGT GCCGCAGTAC TCCATGGTGC AGAGAGCTTG CTCCTGTGGA GGAAGTGTCT

781 ATGTGGTCCC CAGCTCCTCT GTCTGCCTGT CCACTGAGGG GCACCCATGG CTCAGCAGAA

841 GGGCTATTCT TGGGGTTCCC GGTCCTCCTC CAGCCCCGCT AATCTGTGTA GGCCTCAAGT

901 GCTGTGTGTT TGTAAGCATT GTCATCCACA GTCCTATTGT ACGAGCTGGT TCACCCGCAG

961 CTCTGAGCTG CTCTCCAGCC CCAGCCCTTT CTTCCTGTGC CCCTACCCCC GCTGGGATGA

1021 CTCTCCTCAC CCTCCCTGGG GCGACAACCG CCCTGTCTGT AATGAGTGGC AGTCCCAAGC

1081 TTCCTGACTG GCTTCCGCAG CTCTCTGACT CCCCTAAACA AGGCCTCAGG GACTCCACAT

1141 CCAAATTAAG GCGGCACCTG GTGGCAGGTT GGCATTTTCC GGTGTCCTAT CTATGAAAGA

1201 CAGGAAGACA GCTGGGAGCA AACTCCCCTG GGCCAGACTC TTGGAAACAT AAAGGCTTGG

1261 GTGCCCAGCT GGGGACCGGG AGAAAGTCTA AAACACGGGA CTGGGCCAAG GACCCCACAG

1321 GTCCCTGTCT CATTAGGTCC CCTGAAACGT GTGGAAGCTA AAATGGCATT CACGTGATTC

1381 TTGATCATTT AACAGTGGAT TCTGATCTGA TACTACACTG AGAAGTGCCC CTGGGCCGGG

1441 CGCGGTGGCT CACGCCTGTA ATCCCAGCAC TTTGGGAGGC CGAGGCGGGC GGATCACAAG

1501 GAGATTGAGA CCATCCTGGC TAACACGGTG AAACCCTGTC TCTACTAAAA ATACAAAAAA

1561 TTAGCCAGGC ATGGTGGCAG GCGCCTCTAG TACTAGCTAC TCGGGAGGCT GAGGCAGGAG

1621 AATGGTGTGA ACCCGGGAGG CGGAACTTGC AGTGAGCCAA GATTGTGCCA CTGCACTCTA

1681 GCATGGGCGA CAGAGCAAGA CTCAGTCTCA AAAAAAAAAA AAAAAAAAAA AAAAAAAAA

B: aminoacid sequence (SEQ ID NO:14) length: 150 amino acid

1 MVQRACSCGG SVYVVPSSSV CLSTEGHPWL SRRAILGVPG PPPAPLICVG LKCCVFVSIV

61 IHSPIVRAGS PAALSCSPAP ALSSCAPTPA GMTLLTLPGA TTALSVMSGS PKLPDWLPQL

121 SDSPKQGLRD STSKLRRHLV AGWHFPVSYL

C. Nucleotide and amino acid composite sequence (SEQ ID NO:15)

Clone number: PP10199

Start code: 744ATG stops coding: 1194TGA

Protein molecular weight: 15496.34

1 GT TCC AGA GCC ACT TTT AAG ATT CTT CAA TTC CAA ATG CAT GTC TTT 47

48 TTT TAA AAA AAA GAA AGA AAG AAA AAT AAG TTT CTA ATA TTA GAG AAG 95

96 TAC AGC CCT GAA TTG GGT TTT GTG TCC ACT GCT GGA CCC CAT GAG GGC 143

144 CAG GTG GAG TGG ACC TCT GCA GCC CCA GTT GTG TGC ACT CTC TGT TTG 191

192 GTG CAA ATT CCA GTT TGC TGG TTC TCA ATA GCA AGA CCA GCC TGA GAC 239

240 CAC CTG TCC TGC TCT TCC CAT GAG AGG GCC GAA TGC TCC CAG CCT CCA 287

288 TGC CAT GTC CTG TTC CTG GGG TCC TGG GGG TCA TTG CAG CCT GTA TGT 335

336 GCT TCC TCC AGC CAG GGT GAT CAT CGG GTG CCC CAG TGA GCC CCA GCA 383

384 CTG AGG GTC AGC CCC AGG CAC TGT CAA AGG TGA GAG CTC AGA GGC TGT 431

432 GCC CAG AAA GAG AGG TGG GCC CTG CCT GCC CTG GAC GGA GGG AGA GAG 479

480 GCT TCT CAG AGC CCG AGG CAT GAA CCC TCA GGT GGG TCG TGG CCA TAG 527

528 TCA GAT GAT GGC TGC TGG TGA GCT CAG TGA CCA GGC GTC TTC AGG CAG 575

576 CTC ATA AGT TTG AGA GGA CAC AGC CTA AGG GAG GTT TGC TGG GGA GTA 623

624 GCC CCA CTT CCA CCC TGA ATA GAC AAG AGA TGG TAA AGC AGG TAC CCA 671

672 GCA CTT AGT GCT TTC TTG GGG ATA TCG CGT GGG TCC CCG GGG GCC TGG 719

720 GTG CCC GAA GTG CCG CAG TAC TCC ATG GTG CAG AGA GCT TGC TCC TGT 767

1 Met Val Gln Arg Ala Cys Ser Cys 8

768 GGA GGA AGT GTC TAT GTG GTC CCC AGC TCC TCT GTC TGC CTG TCC ACT 815

9 Gly Gly Ser Val Tyr Val Val Pro Ser Ser Ser Val Cys Leu Ser Thr 24

816 GAG GGG CAC CCA TGG CTC AGC AGA AGG GCT ATT CTT GGG GTT CCC GGT 863

25 Glu Gly His Pro Trp Leu Ser Arg Arg Ala Ile Leu Gly Val Pro Gly 40

864 CCT CCT CCA GCC CCG CTA ATC TGT GTA GGC CTC AAG TGC TGT GTG TTT 911

41 Pro Pro Pro Ala Pro Leu Ile Cys Val Gly Leu Lys Cys Cys Val Phe 56

912 GTA AGC ATT GTC ATC CAC AGT CCT ATT GTA CGA GCT GGT TCA CCC GCA 959

57 Val Ser Ile Val Ile His Ser Pro Ile Val Arg Ala Gly Ser Pro Ala 72

960 GCT CTG AGC TGC TCT CCA GCC CCA GCC CTT TCT TCC TGT GCC CCT ACC 1007

73 Ala Leu Ser Cys Ser Pro Ala Pro Ala Leu Ser Ser Cys Ala Pro Thr 88

1008 CCC GCT GGG ATG ACT CTC CTC ACC CTC CCT GGG GCG ACA ACC GCC CTG 1055

89 Pro Ala Gly Met Thr Leu Leu Thr Leu Pro Gly Ala Thr Thr Ala Leu 104

1056 TCT GTA ATG AGT GGC AGT CCC AAG CTT CCT GAC TGG CTT CCG CAG CTC 1103

105 Ser Val Met Ser Gly Ser Pro Lys Leu Pro Asp Trp Leu Pro Gln Leu 120

1104 TCT GAC TCC CCT AAA CAA GGC CTC AGG GAC TCC ACA TCC AAA TTA AGG 1151

121 Ser Asp Ser Pro Lys Gln Gly Leu Arg Asp Ser Thr Ser Lys Leu Arg 136

1152 CGG CAC CTG GTG GCA GGT TGG CAT TTT CCG GTG TCC TAT CTA TGA AAG 1199

137 Arg His Leu Val Ala Gly Trp His Phe Pro Val Ser Tyr Leu *** 151

1200 ACA GGA AGA CAG CTG GGA GCA AAC TCC CCT GGG CCA GAC TCT TGG AAA 1247

1248 CAT AAA GGC TTG GGT GCC CAG CTG GGG ACC GGG AGA AAG TCT AAA ACA 1295

1296 CGG GAC TGG GCC AAG GAC CCC ACA GGT CCC TGT CTC ATT AGG TCC CCT 1343

1344 GAA ACG TGT GGA AGC TAA AAT GGC ATT CAC GTG ATT CTT GAT CAT TTA 1391

1392 ACA GTG GAT TCT GAT CTG ATA CTA CAC TGA GAA GTG CCC CTG GGC CGG 1439

1440 GCG CGG TGG CTC ACG CCT GTA ATC CCA GCA CTT TGG GAG GCC GAG GCG 1487

1488 GGC GGA TCA CAA GGA GAT TGA GAC CAT CCT GGC TAA CAC GGT GAA ACC 1535

1536 CTG TCT CTA CTA AAA ATA CAA AAA ATT AGC CAG GCA TGG TGG CAG GCG 1583

1584 CCT CTA GTA CTA GCT ACT CGG GAG GCT GAG GCA GGA GAA TGG TGT GAA 1631

1632 CCC GGG AGG CGG AAC TTG CAG TGA GCC AAG ATT GTG CCA CTG CAC TCT 1679

1680 AGC ATG GGC GAC AGA GCA AGA CTC AGT CTC AAA AAA AAA AAA AAA AAA 1727

1728 AAA AAA AAA AAA 1739

6.PP10226

A: nucleotide sequence (SEQ ID NO:16) length: 1012bp

1 GTGAGAGAGG GGTTTGGAAA TACCAGACTA TAATTGTGGA TTTGTCCATT ACTCCTTTCA

61 GTTCTAGCAG TTTTTGCTTC TTGTGTTTTG AAGCTCTGTT ATTTGATAAA AATTTTTAGA

121 ATTTTTAATG TTTATTTTAG AATGTATAAA ATTTTAGAAT TTATATGGAT AAATTGAATC

181 CTCTATCATT ATAACATTAT GTTCTTTATG CCTGTAATAT TTTTTGCTGC AAAATCTACT

241 GTCTTAAATA ATATAGACAC AACAGCCTGA TTAGTGTTTG CATAGTACAT CTTCCCCTTC

301 TTCCATTGTT TTACATTTAG CCTATTTGTG CTTTAAAAAA ATTTAAGTAC CTATATTGTA

361 GGCAGCATAG AGTTGGATCT TGTTTTATTA ATGCACCCTG TTTTGAGAGA GAGAGAGAGA

421 GAGACAGAGA CAGAGACACA GAGAGAGAGT GTGAGCGAGC AAAAGAGATT TATTCTGGTT

481 TTTTTTTGTT TGTTTTTGAG ATGGAGTCTT GCTCTCTTGC TCAGGCTGGA GTGCAGTGGC

541 GCAATCTCAG CTCACTGCAA CCTCCACCTC CTGGGTTCAA GTTATTCTCC TGTCTCAGCC

601 TCCCAAGTAG CTGGGACTAC AGGCCTGTGC CACCATGCCC GGCTACGTTT TGTATTTTTA

661 GTACAGACGG TGTTTCACCA TGTTGGCCAG GCTGGTCTCA AACTCCTGGC CTCAAGTTGA

721 TCTGCTGGCC TCACGCCTGT AATCCTAGTA CTTTGGGAGG CCGAGGCGGG CGGATCTCGA

781 GTTCAGGAGA TCGACCATCC TGGCTAACAC GGTGAAACCT CGTCTCTACT AAAAATACAA

841 AAAATTAGCC GGGCATGGTG GTGGGCACCC GTAGTCCCAG CTACTTGGGA GGCTGAGGCA

901 GGAGAATGGC ATGAATCCAG TAGGCGGAGC TTGCAGTGAG CCAAGATCAC GCCACTGCAC

961 TCCAGCCTGG GTGACAGAGC GAGACTTTGT CTCAAAAAAA AAAAAAAAAA AA

B: aminoacid sequence (SEQ ID NO:17) length: 109 amino acid

1 MHPVLRERER ETETETQRES VSEQKRFILV FFCLFLRWSL ALLLRLECSG AISAHCNLHL

61 LGSSYSPVSA SQVAGTTGLC HHARLRFVFL VQTVFHHVGQ AGLKLLASS

C. Nucleotide and amino acid composite sequence (SEQ ID NO:18)

Clone number: PP10226

Start code: 391ATG stops coding: 718TGA

Protein molecular weight: 12262.59

1 GTG AGA GAG GGG TTT GGA AAT ACC AGA CTA TAA TTG TGG ATT TGT CCA 48

49 TTA CTC CTT TCA GTT CTA GCA GTT TTT GCT TCT TGT GTT TTG AAG CTC 96

97 TGT TAT TTG ATA AAA ATT TTT AGA ATT TTT AAT GTT TAT TTT AGA ATG 144

145 TAT AAA ATT TTA GAA TTT ATA TGG ATA AAT TGA ATC CTC TAT CAT TAT 192

193 AAC ATT ATG TTC TTT ATG CCT GTA ATA TTT TTT GCT GCA AAA TCT ACT 240

241 GTC TTA AAT AAT ATA GAC ACA ACA GCC TGA TTA GTG TTT GCA TAG TAC 288

289 ATC TTC CCC TTC TTC CAT TGT TTT ACA TTT AGC CTA TTT GTG CTT TAA 336

337 AAA AAT TTA AGT ACC TAT ATT GTA GGC AGC ATA GAG TTG GAT CTT GTT 384

385 TTA TTA ATG CAC CCT GTT TTG AGA GAG AGA GAG AGA GAG ACA GAG ACA 432

1 Met His Pro Val Leu Arg Glu Arg Glu Arg Glu Thr Glu Thr 14

433 GAG ACA CAG AGA GAG AGT GTG AGC GAG CAA AAG AGA TTT ATT CTG GTT 480

15 Glu Thr Gln Arg Glu Ser Val Ser Glu Gln Lys Arg Phe Ile Leu Val 30

481 TTT TTT TGT TTG TTT TTG AGA TGG AGT CTT GCT CTC TTG CTC AGG CTG 528

31 Phe Phe Cys Leu Phe Leu Arg Trp Ser Leu Ala Leu Leu Leu Arg Leu 46

529 GAG TGC AGT GGC GCA ATC TCA GCT CAC TGC AAC CTC CAC CTC CTG GGT 576

47 Glu Cys Ser Gly Ala Ile Ser Ala His Cys Asn Leu His Leu Leu Gly 62

577 TCA AGT TAT TCT CCT GTC TCA GCC TCC CAA GTA GCT GGG ACT ACA GGC 624

63 Ser Ser Tyr Ser Pro Val Ser Ala Ser Gln Val Ala Gly Thr Thr Gly 78

625 CTG TGC CAC CAT GCC CGG CTA CGT TTT GTA TTT TTA GTA CAG ACG GTG 672

79 Leu Cys His His Ala Arg Leu Arg Phe Val Phe Leu Val Gln Thr Val 94

673 TTT CAC CAT GTT GGC CAG GCT GGT CTC AAA CTC CTG GCC TCA AGT TGA 720

95 Phe His His Val Gly Gln Ala Gly Leu Lys Leu Leu Ala Ser Ser *** 110

721 TCT GCT GGC CTC ACG CCT GTA ATC CTA GTA CTT TGG GAG GCC GAG GCG 768

769 GGC GGA TCT CGA GTT CAG GAG ATC GAC CAT CCT GGC TAA CAC GGT GAA 816

817 ACC TCG TCT CTA CTA AAA ATA CAA AAA ATT AGC CGG GCA TGG TGG TGG 864

865 GCA CCC GTA GTC CCA GCT ACT TGG GAG GCT GAG GCA GGA GAA TGG CAT 912

913 GAA TCC AGT AGG CGG AGC TTG CAG TGA GCC AAG ATC ACG CCA CTG CAC 960

961 TCC AGC CTG GGT GAC AGA GCG AGA CTT TGT CTC AAA AAA AAA AAA AAA 1008

1009 AAA A 1012

7.SP2114a

A: nucleotide sequence (SEQ ID NO:19) length: 2546bp

1 GGCCAGTCAA GATGGCCGCC GCTGGGTGAG GCAAGCTGGC GCGCCGCGGG GGCGTCTGGG

61 AGTTGTAGTT CGGGACGGCG GGCTGACGCA CTTCGCCGCC GGCCGACGGG CGCCATTGTG

121 CGGCGCGCGC CGGGACTCTG CCCACTTCCA CCAGAGACAC ATTGAGAAGG AGGAAACTAT

181 GGCCTCCAGG CTTCCGACGG CCTGGTCCTG TGAACCAGAG ACCTTTGAAG ATGTAACACT

241 GGGTTTTACC CCGGAAGAGT GGGGACTGCT GGACCTCAAA CAGAAGTCCC TGTACAGGGA

301 AGTGATGCTG GAGAACTACA GGAACCTGGT CTCAGTGGAA CATCAGCTTT CCAAACCAGA

361 TGTGGTATCT CAGTTAGAGG AGGCAGAAGA TTTCTGGCCA GTGGAGAGAG GAATTCCTCA

421 AGACACCATT CCTGAGTATC CTGAGCTCCA GCTGGACCCT AAATTGGATC CTCTTCCTGC

481 TGAGAGTCCC CTAATGAACA TTGAGGTTGT TGAGGTCCTC ACACTGAACC AGGAGGTGGC

541 TGGTCCCCGG AATGCCCAGA TCCAGGCCCT ATATGCTGAA GATGGAAGCC TGAGTGCAGA

601 TGCCCCCAGT GAGCAGATCC AACAGCAGGG CAAGCATCCA GGTGACCCTG AGGCCGCGCG

661 CCAGAGGTTC CGGCAGTTCC GTTATAAGGA CATGACAGGT CCCCGGGAGG CCCTGGACCA

721 GCTCCGAGAG CTGTGTCACC AGTGGCTACA GCCTAAGGCA CGCTCCAAGG AGCAGATCCT

781 GGAGCTGCTG GTGCTGGAGC AGTTCCTAGG TACACTGCCT GTGAAGCTCC GGACATGGGT

841 GGAATCGCAG CACCCAGAGA ACTGCCAAGA GGTGGTGGCC CTGGTAGAGG GTGTGACCTG

901 GATGTCTGAG GAGGAAGTAC TTCCTGCAGG ACAACCTGCC GAGGGCACCA CCTGCTGCCT

961 CGAGGTCACT GCCCAGCAGG AGGAGAAGCA GGAGGATGCA GCCATCTGCC CAGTGACAGT

1021 GCTCCCTGAG GAGCCAGTGA CCTTCCAGGA TGTGGCTGTG GACTTCAGCC GGGAGGAGTG

1081 GGGGCTGCTG GGCCCGACAC AGAGGACCGA GTACCGCGAT GTGATGCTGG AGACCTTTGG

1141 GCACCTGGTC TCTGTGGGGT GGGAGACTAC ACTGGAAAAT AAAGAGTTAG CTCCAAATTC

1201 TGACATTCCT GAGGAAGAAC CAGCCCCCAG CCTGAAAGTA CAAGAATCCT CAAGGGATTG

1261 TGCCTTGTCC TCTACATTAG AAGATACCTT GCAGGGTGGG GTCCAGGAAG TCCAAGACAC

1321 AGTGTTGAAG CAGATGGAGT CTGCTCAGGA AAAAGACCTT CCTCAGAAGA AGCACTTTGA

1381 CAACCGTGAG TCCCAGGCAA ACAGTGGTGC TCTTGACACA AACCAAGTTT CGCTCCAGAA

1441 AATTGACAAC CCTGAGTCCC AGGCAAACAG TGGCGCTCTT GACACAAACC AAGTTTTGCT

1501 CCACAAAATT CCTCCTAGAA AACGATTGCG CAAACGTGAC TCACAAGTTA AAAGTATGAA

1561 ACATAATTCA CGTGTAAAAA TTCATCAGAA GAGCTGTGAA AGGCAAAAGG CCAAGGAAGG

1621 CAATGGTTGT AGGAAAACCT TCAGTCGGAG TACTAAACAG ATTACGTTTA TAAGAATTCA

1681 CAAGGGGAGC CAAGTTTGCC GATGCAGTGA ATGTGGTAAA ATATTCCGGA ACCCAAGATA

1741 CTTTTCTGTG CATAAGAAAA TCCATACCGG AGAGAGGCCC TATGTGTGTC AAGACTGTGG

1801 GAAAGGATTT GTTCAGAGCT CTTCCCTCAC ACAGCATCAG AGAGTTCATT CTGGAGAGAG

1861 ACCATTTGAA TGTCAGGAGT GTGGGAGGAC CTTCAATGAT CGCTCAGCCA TCTCCCAGCA

1921 CCTGAGGACT CACACTGGCG CTAAGCCCTA CAAGTGTCAG GACTGTGGAA AAGCCTTCCG

1981 CCAGAGTTCC CACCTCATCA GACATCAGAG GACTCACACC GGGGAGCGCC CATATGCATG

2041 CAACAAATGT GGAAAGGCCT TCACCCAGAG CTCACACCTT ATTGGGCACC AGAGAACCCA

2101 CAATAGGACA AAGCGAAAGA AGAAACAGCC TACCTCATAG CTCTCAAGCC AGTTGAAGAA

2161 ACCTTGCCTT TTCAGCTTGA CCCTGCAATA TAACATGCAC AGGCCTGCTT GTGAATCAGG

2221 ACTGAATGTG AAAGGGAAGT ATTGAGTGAG GACATTCCCA AAACCAAAGG ACAACTGAGG

2281 AGACTGCCCA GCACATAATG AATAAATAAG AAAATGAGTG AGGAGTTATT AACATCATTT

2341 GGAAAAAAGA TTTCCCATTC ACTTGATATT GTTTGTTCAC TCATTTAGTC ATTAAAAGTG

2401 AGATTAATAA AATCTGAAAA TGTTATATAA TAACTTTAAA AAGCCAGGTA ATTAATAATC

2461 TGCACTGATA TTACATCCAC AGTACCACAG TATTTATGTG TATGAATTAA GGATTAAAAG

2521 ATAATGTGGA TAAAAAAAAA AAAAAA

B: aminoacid sequence (SEQ ID NO:20) length: 653 amino acid

1 MASRLPTAWS CEPETFEDVT LGFTPEEWGL LDLKQKSLYR EVMLENYRNL VSVEHQLSKP

61 DVVSQLEEAE DFWPVERGIP QDTIPEYPEL QLDPKLDPLP AESPLMNIEV VEVLTLNQEV

121 AGPRNAQIQA LYAEDGSLSA DAPSEQIQQQ GKHPGDPEAA RQRFRQFRYK DMTGPREALD

181 QLRELCHQWL QPKARSKEQI LELLVLEQFL GTLPVKLRTW VESQHPENCQ EVVALVEGVT

241 WMSEEEVLPA GQPAEGTTCC LEVTAQQEEK QEDAAICPVT VLPEEPVTFQ DVAVDFSREE

301 WGLLGPTQRT EYRDVMLETF GHLVSVGWET TLENKELAPN SDIPEEEPAP SLKVQESSRD

361 CALSSTLEDT LQGGVQEVQD TVLKQMESAQ EKDLPQKKHF DNRESQANSG ALDTNQVSLQ

421 KIDNPESQAN SGALDTNQVL LHKIPPRKRL RKRDSQVKSM KHNSRVKIHQ KSCERQKAKE

481 GNGCRKTFSR STKQITFIRI HKGSQVCRCS ECGKIFRNPR YFSVHKKIHT GERPYVCQDC

541 GKGFVQSSSL TQHQRVHSGE RPFECQECGR TFNDRSAISQ HLRTHTGAKP YKCQDCGKAF

601 RQSSHLIRHQ RTHTGERPYA CNKCGKAFTQ SSHLIGHQRT HNRTKRKKKQ PTS

C. Nucleotide and amino acid composite sequence (SEQ ID NO:21)

Clone number: SP2114a

Start code: 179ATG stops coding: 2138TAG

Protein molecular weight: 74247.90

1 G GCC AGT CAA GAT GGC CGC CGC TGG GTG AGG CAA GCT GGC GCG CCG 46

47 CGG GGG CGT CTG GGA GTT GTA GTT CGG GAC GGC GGG CTG ACG CAC TTC 94

95 GCC GCC GGC CGA CGG GCG CCA TTG TGC GGC GCG CGC CGG GAC TCT GCC 142

143 CAC TTC CAC CAG AGA CAC ATT GAG AAG GAG GAA ACT ATG GCC TCC AGG 190

1 Met Ala Ser Arg 4

191 CTT CCG ACG GCC TGG TCC TGT GAA CCA GAG ACC TTT GAA GAT GTA ACA 238

5 Leu Pro Thr Ala Trp Ser Cys Glu Pro Glu Thr Phe Glu Asp Val Thr 20

239 CTG GGT TTT ACC CCG GAA GAG TGG GGA CTG CTG GAC CTC AAA CAG AAG 286

21 Leu Gly Phe Thr Pro Glu Glu Trp Gly Leu Leu Asp Leu Lys Gln Lys 36

287 TCC CTG TAC AGG GAA GTG ATG CTG GAG AAC TAC AGG AAC CTG GTC TCA 334

37 Ser Leu Tyr Arg Glu Val Met Leu Glu Asn Tyr Arg Asn Leu Val Ser 52

335 GTG GAA CAT CAG CTT TCC AAA CCA GAT GTG GTA TCT CAG TTA GAG GAG 382

53 Val Glu His Gln Leu Ser Lys Pro Asp Val Val Ser Gln Leu Glu Glu 68

383 GCA GAA GAT TTC TGG CCA GTG GAG AGA GGA ATT CCT CAA GAC ACC ATT 430

69 Ala Glu Asp Phe Trp Pro Val Glu Arg Gly Ile Pro Gln Asp Thr Ile 84

431 CCT GAG TAT CCT GAG CTC CAG CTG GAC CCT AAA TTG GAT CCT CTT CCT 478

85 Pro Glu Tyr Pro Glu Leu Gln Leu Asp Pro Lys Leu Asp Pro Leu Pro 100

479 GCT GAG AGT CCC CTA ATG AAC ATT GAG GTT GTT GAG GTC CTC ACA CTG 526

101 Ala Glu Ser Pro Leu Met Asn Ile Glu Val Val Glu Val Leu Thr Leu 116

527 AAC CAG GAG GTG GCT GGT CCC CGG AAT GCC CAG ATC CAG GCC CTA TAT 574

117 Asn Gln Glu Val Ala Gly Pro Arg Asn Ala Gln Ile Gln Ala Leu Tyr 132

575 GCT GAA GAT GGA AGC CTG AGT GCA GAT GCC CCC AGT GAG CAG ATC CAA 622

133 Ala Glu Asp Gly Ser Leu Ser Ala Asp Ala Pro Ser Glu Gln Ile Gln 148

623 CAG CAG GGC AAG CAT CCA GGT GAC CCT GAG GCC GCG CGC CAG AGG TTC 670

149 Gln Gln Gly Lys His Pro Gly Asp Pro Glu Ala Ala Arg Gln Arg Phe 164

671 CGG CAG TTC CGT TAT AAG GAC ATG ACA GGT CCC CGG GAG GCC CTG GAC 718

165 Arg Gln Phe Arg Tyr Lys Asp Met Thr Gly Pro Arg Glu Ala Leu Asp 180

719 CAG CTC CGA GAG CTG TGT CAC CAG TGG CTA CAG CCT AAG GCA CGC TCC 766

181 Gln Leu Arg Glu Leu Cys His Gln Trp Leu Gln Pro Lys Ala Arg Ser 196

767 AAG GAG CAG ATC CTG GAG CTG CTG GTG CTG GAG CAG TTC CTA GGT ACA 814

197 Lys Glu Gln Ile Leu Glu Leu Leu Val Leu Glu Gln Phe Leu Gly Thr 212

815 CTG CCT GTG AAG CTC CGG ACA TGG GTG GAA TCG CAG CAC CCA GAG AAC 862

213 Leu Pro Val Lys Leu Arg Thr Trp Val Glu Ser Gln His Pro Glu Asn 228

863 TGC CAA GAG GTG GTG GCC CTG GTA GAG GGT GTG ACC TGG ATG TCT GAG 910

229 Cys Gln Glu Val Val Ala Leu Val Glu Gly Val Thr Trp Met Ser Glu 244

911 GAG GAA GTA CTT CCT GCA GGA CAA CCT GCC GAG GGC ACC ACC TGC TGC 958

245 Glu Glu Val Leu Pro Ala Gly Gln Pro Ala Glu Gly Thr Thr Cys Cys 260

959 CTC GAG GTC ACT GCC CAG CAG GAG GAG AAG CAG GAG GAT GCA GCC ATC 1006

261 Leu Glu Val Thr Ala Gln Gln Glu Glu Lys Gln Glu Asp Ala Ala Ile 276

1007 TGC CCA GTG ACA GTG CTC CCT GAG GAG CCA GTG ACC TTC CAG GAT GTG 1054

277 Cys Pro Val Thr Val Leu Pro Glu Glu Pro Val Thr Phe Gln Asp Val 292

1055 GCT GTG GAC TTC AGC CGG GAG GAG TGG GGG CTG CTG GGC CCG ACA CAG 1102

293 Ala Val Asp Phe Ser Arg Glu Glu Trp Gly Leu Leu Gly Pro Thr Gln 308

1103 AGG ACC GAG TAC CGC GAT GTG ATG CTG GAG ACC TTT GGG CAC CTG GTC 1150

309 Arg Thr Glu Tyr Arg Asp Val Met Leu Glu Thr Phe Gly His Leu Val 324

1151 TCT GTG GGG TGG GAG ACT ACA CTG GAA AAT AAA GAG TTA GCT CCA AAT 1198

325 Ser Val Gly Trp Glu Thr Thr Leu Glu Asn Lys Glu Leu Ala Pro Asn 340

1199 TCT GAC ATT CCT GAG GAA GAA CCA GCC CCC AGC CTG AAA GTA CAA GAA 1246

341 Ser Asp Ile Pro Glu Glu Glu Pro Ala Pro Ser Leu Lys Val Gln Glu 356

1247 TCC TCA AGG GAT TGT GCC TTG TCC TCT ACA TTA GAA GAT ACC TTG CAG 1294

357 Ser Ser Arg Asp Cys Ala Leu Ser Ser Thr Leu Glu Asp Thr Leu Gln 372

1295 GGT GGG GTC CAG GAA GTC CAA GAC ACA GTG TTG AAG CAG ATG GAG TCT 1342

373 Gly Gly Val Gln Glu Val Gln Asp Thr Val Leu Lys Gln Met Glu Ser 388

1343 GCT CAG GAA AAA GAC CTT CCT CAG AAG AAG CAC TTT GAC AAC CGT GAG 1390

389 Ala Gln Glu Lys Asp Leu Pro Gln Lys Lys His Phe Asp Asn Arg Glu 404

1391 TCC CAG GCA AAC AGT GGT GCT CTT GAC ACA AAC CAA GTT TCG CTC CAG 1438

405 Ser Gln Ala Asn Ser Gly Ala Leu Asp Thr Asn Gln Val Ser Leu Gln 420

1439 AAA ATT GAC AAC CCT GAG TCC CAG GCA AAC AGT GGC GCT CTT GAC ACA 1486

421 Lys Ile Asp Asn Pro Glu Ser Gln Ala Asn Ser Gly Ala Leu Asp Thr 436

1487 AAC CAA GTT TTG CTC CAC AAA ATT CCT CCT AGA AAA CGA TTG CGC AAA 1534

437 Asn Gln Val Leu Leu His Lys Ile Pro Pro Arg Lys Arg Leu Arg Lys 452

1535 CGT GAC TCA CAA GTT AAA AGT ATG AAA CAT AAT TCA CGT GTA AAA ATT 1582

453 Arg Asp Ser Gln Val Lys Ser Met Lys His Asn Ser Arg Val Lys Ile 468

1583 CAT CAG AAG AGC TGT GAA AGG CAA AAG GCC AAG GAA GGC AAT GGT TGT 1630

469 His Gln Lys Ser Cys Glu Arg Gln Lys Ala Lys Glu Gly Asn Gly Cys 484

1631 AGG AAA ACC TTC AGT CGG AGT AGT AAA CAG ATT ACG TTT ATA AGA ATT 1678

485 Arg Lys Thr Phe Ser Arg Ser Thr Lys Gln Ile Thr Phe Ile Arg Ile 500

1679 CAC AAG GGG AGC CAA GTT TGC CGA TGC AGT GAA TGT GGT AAA ATA TTC 1726

501 His Lys Gly Ser Gln Val Cys Arg Cys Ser Glu Cys Gly Lys Ile Phe 516

1727 CGG AAC CCA AGA TAC TTT TCT GTG CAT AAG AAA ATC CAT ACC GGA GAG 1774

517 Arg Asn Pro Arg Tyr Phe Ser Val His Lys Lys Ile His Thr Gly Glu 532

1775 AGG CCC TAT GTG TGT CAA GAC TGT GGG AAA GGA TTT GTT CAG AGC TCT 1822

533 Arg Pro Tyr Val Cys Gln Asp Cys Gly Lys Gly Phe Val Gln Ser Ser 548

1823 TCC CTC ACA CAG CAT CAG AGA GTT CAT TCT GGA GAG AGA CCA TTT GAA 1870

549 Ser Leu Thr Gln His Gln Arg Val His Ser Gly Glu Arg Pro Phe Glu 564

1871 TGT CAG GAG TGT GGG AGG ACC TTC AAT GAT CGC TCA GCC ATC TCC CAG 1918

565 Cys Gln Glu Cys Gly Arg Thr Phe Asn Asp Arg Ser Ala Ile Ser Gln 580

1919 CAC CTG AGG ACT CAC ACT GGC GCT AAG CCC TAC AAG TGT CAG GAC TGT 1966

581 His Leu Arg Thr His Thr Gly Ala Lys Pro Tyr Lys Cys Gln Asp Cys 596

1967 GGA AAA GCC TTC CGC CAG AGT TCC CAC CTC ATC AGA CAT CAG AGG ACT 2014

597 Gly Lys Ala Phe Arg Gln Ser Ser His Leu Ile Arg His Gln Arg Thr 612

2015 CAC ACC GGG GAG CGC CCA TAT GCA TGC AAC AAA TGT GGA AAG GCC TTC 2062

613 His Thr Gly Glu Arg Pro Tyr Ala Cys Asn Lys Cys Gly Lys Ala Phe 628

2063 ACC CAG AGC TCA CAC CTT ATT GGG CAC CAG AGA ACC CAC AAT AGG ACA 2110

629 Thr Gln Ser Ser His Leu Ile Gly His Gln Arg Thr His Asn Arg Thr 644

2111 AAG CGA AAG AAG AAA CAG CCT ACC TCA TAG CTC TCA AGC CAG TTG AAG 2158

645 Lys Arg Lys Lys Lys Gln Pro Thr Ser *** 654

2159 AAA CCT TGC CTT TTC AGC TTG ACC CTG CAA TAT AAC ATG CAC AGG CCT 2206

2207 GCT TGT GAA TCA GGA CTG AAT GTG AAA GGG AAG TAT TGA GTG AGG ACA 2254

2255 TTC CCA AAA CCA AAG GAC AAC TGA GGA GAC TGC CCA GCA CAT AAT GAA 2302

2303 TAA ATA AGA AAA TGA GTG AGG AGT TAT TAA CAT CAT TTG GAA AAA AGA 2350

2351 TTT CCC ATT CAC TTG ATA TTG TTT GTT CAC TCA TTT AGT CAT TAA AAG 2398

2399 TGA GAT TAA TAA AAT CTG AAA ATG TTA TAT AAT AAC TTT AAA AAG CCA 2446

2447 GGT AAT TAA TAA TCT GCA CTG ATA TTA CAT CCA CAG TAC CAC AGT ATT 2494

2495 TAT GTG TAT GAA TTA AGG ATT AAA AGA TAA TGT GGA TAA AAA AAA AAA 2542

2543 AAA A 2546

All quote in this application as a reference at all documents that the present invention mentions, just quoted as a reference separately as each piece document.Should be understood that in addition those skilled in the art can make various changes or modifications the present invention after having read above-mentioned teachings of the present invention, these equivalent form of values fall within the application's appended claims institute restricted portion equally.

Claims (10)

1. isolating people's albumen with cancer suppressing function, it is characterized in that it comprises the polypeptide of the aminoacid sequence with the group of being selected from down: SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:11, SEQ ID NO:14, SEQ ID NO:17, SEQ ID NO:20.

2. albumen as claimed in claim 1, it is characterized in that this proteic aminoacid sequence is selected from down group: SEQ ID NO:2, SEQ ID NO:5, SEQ ID NO:8, SEQ ID NO:11, SEQ ID NO:14, SEQ ID NO:17, SEQ ID NO:20.

3. isolating polynucleotide is characterized in that, it comprises a nucleotide sequence, and this nucleotide sequence is selected from down group:

(a) the proteic according to claim 1 polynucleotide of coding;

(b) with polynucleotide (a) complementary polynucleotide.

4. polynucleotide as claimed in claim 3 is characterized in that, the sequence of these polynucleotide is selected from down group:

The coding region sequence of SEQ ID NO:3, SEQ ID NO:6, SEQ ID NO:9, SEQ ID NO:12, SEQ ID NO:15, SEQ ID NO:18, SEQ ID NO:21.

5. polynucleotide as claimed in claim 3 is characterized in that, the sequence of these polynucleotide is selected from down group:

The full length sequence of SEQ ID NO:3, SEQ ID NO:6, SEQ ID NO:9, SEQ ID NO:12, SEQ ID NO:15, SEQ ID NO:18, SEQ ID NO:21.

6. a carrier is characterized in that, it contains the described polynucleotide of claim 3.

7. a genetically engineered host cell is characterized in that, it is a kind of host cell that is selected from down group:

(a) host cell that transforms or transduce with the described carrier of claim 6;

(b) host cell that transforms or transduce with the described polynucleotide of claim 3.

8. the preparation method of the polypeptide of the people's protein-active with cancer suppressing function is characterized in that this method comprises:

(a) have under the proteic condition of people of cancer suppressing function suitable the expression, cultivate the described host cell of claim 7;

(b) from culture, isolate the polypeptide of people's protein-active with cancer suppressing function.

9. energy and the people's protein-specific bonded antibody with cancer suppressing function, wherein said people's albumen with cancer suppressing function has the aminoacid sequence of the group of being selected from down: SEQ ID NO:2, SEQ ID NO:8, SEQ ID NO:11, SEQ IDNO:14, SEQ ID NO:17.

10. a pharmaceutical composition is characterized in that, it contains the described albumen of claim 1 and the pharmaceutically acceptable carrier of safe and effective amount.