CN1257922A - Cytokine signal transfer arrestin IV - Google Patents

Abstract

The present invention discloses a new polynucleotide, the polypeptide coded with said polynucleotide and the process for preparing the polypeptide using said polynucleotide. Said polypeptide is a member in human cytokine inducible SH2-containing protein (CIS) family.

Description

Cytokine signal transfer arrestin IV

The present invention relates to a kind of new polynucleotide,, and utilize described polynucleotide to produce the method for polypeptide by the polypeptide of this polynucleotide encoding.In particular, polypeptide of the present invention is a member of people CIS (Cytokine inducible SH2-containing protein) protein family.

Cytokine has important regulation to functions such as the propagation of mammalian cell, differentiation, growths.Cytokine can be passed to its information transmission in the nucleus by the JAK-STAT approach, thereby induces, regulates Expression of Related Genes to bring into play its function (Silvennoinen.O.et al. (1997) APMIS.105,497-509; Schindler, C. (1995) Annu RevBiochem 64,621-651; James E., Darnell Jr. (1997) Science 277,1630-1635).The CIS family member is proved to be has regulating effect to JAK-STAT signal pipeline.

Obviously, the function of JAK-STAT approach is inevitable directly related with the receptor-mediated biologically of relevant cell factor, yet the relevant the most direct evidence of its function comes from the research to this approach natural defect patient and gene knockout or transgenation experiment.For example, all show as T cell, NK cell shortage and B cell function defective on X-SCID (X-linked severe combined immunodeficiency) patient and the SCID patients clinical, their pathogeny is thought that respectively the genetic knock-out experiment in mouse has confirmed this conclusion because Jak3 can't activate the sudden change with Jak3; Stat1 is relevant with the natural immunity; Stat2 is relevant with fetal development with Stat3, and their defective all can cause embryonic death; Stat4 participates in the reaction to IL-12, forms relevant with differentiation and the NK cell of Th1; Stat5 is relevant with the lactation function with mammogenesis; Stat6 is then relevant with the reaction to IL-4, and is very important to the formation of the differentiation of Th2 cell, IgE.From above-mentioned evidence as can be seen the JAK-STAT approach exercised important function in vivo.As the attemperator of JAK-STAT approach, CIS also must participate in the regulation and control to all functions of this approach, thereby plays the important physical function in vivo.

The earliest member CIS (cytokine inducible SH2-containingprotein) is in nineteen ninety-five in the CIS gene family, by Yoshimura, the little musculus cdna that A. etc. find when research cytokine induction activatory immediate early gene (immediate early gene).The coded product of this gene can with the IL3 acceptor β chain of phosphorylation or the EPO receptors bind of phosphorylation, and can suppress the cell growth (Yoshimura that IL-3 relies on and EPO relies on, A., et al. (1995) EMBO J.14,2816-2826), and IL3 and EPO can both work by Jak2 and Stat5, so think that CIS albumen may be the negative attemperator that cytokine signaling transmits.1997,---SOCS-1 of mouse (Suppressor OfCytokine Signalling-1) [SOCS-1/JAB/SSI (STAT induced STAT inhibitor)]----is obtained with diverse ways by three different experiments chambers another member of this gene family simultaneously: i) Hilton, D.J. wait by selecting that IL-6 is stimulated unresponsive transformant (Starr, R.et al. (1997) Nature 387,917-921); Ii) Yoshimura, A. etc. with yeast two-hybrid system seek can with Jak2 bonded albumen (Endo, T.A., et al. (1997) Nature 387,921-924); Iii) Kishimoto, T. etc. then be seek to Stat3 have similar antigenic found proteic the time this gene (Naka, T., et al. (1997) Nature 387,924-929).With the SOCS-2 that also has mouse that mouse SOCS-1 announces simultaneously, SOCS-1 gene of-3 genes, rat and people's SOCS-1 (SOCS-1/JAB/SSI-1) gene.

People's SSI-2, SSI-3 gene are very fast also by Kishimoto, T. etc. by the EST homology retrieval to the SH2 structural domain obtain (Minamoto, S., et al. (1997) Biochem BiophysRes Commun 237 (1), 79-83).

Soon, Yoshimura, A. etc. report again by EST retrieval obtain CIS family several newcomers---people's CIS-2～6, wherein CIS-2～4 have been cloned total length.(annotate: wherein, CIS-2 and people SSI-2 only differ from 3 amino acid, and CIS-3 and SSI-3 differ 2, doubt to be same gene.) they these genes together with the genes involved of finding in the past call CIS family (Masuhara, M.Z. (1997) Biochem Biophys Res Commun 239,439-446).

Recently, Hilton, D.J. etc. have obtained 16 genes by database retrieval SOCSbox conserved sequence again, wherein 4 is new SOCS gene (SOCS-4 of mouse, 5,6,7) (HiltonD.J., et al. (1998) Pro Natl Acad Sci.U.S.A.95 (1), 114-119).

An object of the present invention is to provide a kind of new polynucleotide, a newcomer of this polynucleotide encoding CIS family, CIS homologue called after SOCS-4 of the present invention (GenBankAccession No.AF035947).

Another object of the present invention provides a kind of new CIS protein family member, called after SOCS-4.

A further object of the present invention provides a kind of method of utilizing recombinant technology to produce described new CIS protein family member.

Another purpose of the present invention is described polynucleotide and the application of albumen in the medicine of preparation diagnosis and treatment and SOCS-4 relative disease.

The present invention relates to a kind of new isolating polynucleotide,, and utilize described polynucleotide to produce the method for described polypeptide by the polypeptide of this polynucleotide encoding.More particularly, polypeptide of the present invention is a newcomer of CIS protein family.CIS albumen member called after SOCS-4 of the present invention (GenBank Accession No.AF035947).The polynucleotide of the present invention of encoding can be rna form and dna form, and DNA comprises cDNA, genomic dna and synthetic DNA, and DNA can be two strands or strand, if strand can be coding strand or noncoding strand.The present invention also comprised can with nucleic acid fragment, analogue and the derivative of SEQ ID NO:1 hybridization.

In the present invention, term " isolating " is meant that material separates (if natural substance, primal environment promptly is a natural surroundings) from its primal environment.As the polynucleotide under the native state in the active somatic cell and polypeptide be do not have isolating, but same polynucleotide or polypeptide as from native state with in other materials of depositing separately, then be isolating.

In one embodiment of the invention, polynucleotide total length of the present invention is 2069 Nucleotide, and its detailed sequence is seen SEQ ID NO:1, and wherein open reading frame is positioned at 133-933 position Nucleotide.These polynucleotide are so to obtain, with people's placenta λ gt11cDNA library (available from Clontech company) is template, synthetic forward primer 5 '-CCTCCTGCACTGCTGATGCCC-3 ' and reverse primer 5 '-CCTTGCTCTTGCTGGCTCTTCC-3 ', carry out PCR, obtain the purpose fragment of 436bp.This fragment of mark then, with this labeled fragment be probe with amplification after the hybridization of people's placenta λ gt11cDNA library, the picking positive colony after same probe and screening process are carried out multiple sieve to the positive colony that obtains, finally obtains some positive monoclonals.Identify the full length cDNA sequence that obtains SEQ ID NO:1 after checking order.

According to a further aspect in the invention, polynucleotide sequence of the present invention can be used to express or produce the SOCS-4 albumen or the polypeptide of reorganization.In general following steps are arranged: transform proper host cell with the dna fragmentation (or varient) of coding of the present invention SOCS-4 or the recombinant expression vector that contains this dna fragmentation; The host cell of in suitable medium, cultivating; Separation, protein purification from substratum or cell.

The invention still further relates to the expression vector that contains polynucleotide sequence of the present invention and with the genetically engineered host cell of described expression vector.A key character of expression vector is to contain suitable promotor and translation controlling elements.Multiple expression vector can obtain from commercial channels.Carrier commonly used includes, but is not limited to plasmid, phage, clay, Yeast expression carrier, virus, Ti-plasmids etc., but the first-selected plasmid of the most frequently used carrier.Host cell commonly used includes, but is not limited to bacterium, yeast, insect cell, zooblast or vegetable cell etc., but the most frequently used host cell is a bacterium, especially intestinal bacteria.Another expression system commonly used is a mammal cell line, as Hela, and COS clone or Chinese hamster ovary celI system etc.

The known method of skilled person makes up the recombinant expression vector that contains the SOCS-4 encoding sequence in available this area.These methods comprise the extracorporeal recombinant DNA technology, DNA synthetic technology, (J Sambrook etc. 1989, molecular cloning laboratory manuals) such as the interior recombinant technologys of body.

The invention still further relates to antibody, have several different methods to can be used for producing antibody at the SOCS-4 antigenic determinant at polypeptide of the present invention.These antibody include, but is not limited to polyclonal antibody, monoclonal antibody, chimeric antibody, single-chain antibody, the fragment that Fab fragment and Fab expression library produce.

Available SOCS-4 albumen of the production of polyclonal antibody or polypeptide immune animal, as rabbit, mouse, rat etc.Multiple adjuvant can be used for the enhancing immunity reaction, includes but not limited to freund's adjuvant etc.

The SOCS-4 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 etc., PNAS, 1985,81:6851).And the technology of existing manufacture order chain antibody (U.S.Pat No.4946778) also can be used for producing the single-chain antibody of anti-SOCS-4.

According to another aspect of the invention, polynucleotide of the present invention and polypeptide can be used for the diagnosis and the treatment of SOCS-4 relative disease.Known CIS protein family has participated in the adjusting to JAK-STAT signal pipeline.This family comprises other many different members, and different species and individuality also have different members.CIS homologue of the present invention is the newcomer of this family.Therefore the cDNA of the SOCS-4 that is separated to of the present invention can be used to detect unusual that JAK-STAT signal that the defective with CIS homologue of the present invention causes transmits specifically, and the unusual relevant illness of the various and JAK-STAT approach that causes therefrom, and may provide a kind of approach for the treatment of conditions of this class in the future.

Below will further describe the present invention by embodiment, but embodiment only being an illustrative, is not limitation of the present invention.The clone of embodiment 1SOCS-4cDNA and 1. primer amplifications that check order

With people's placenta λ gt11cDNA library (available from Clontech company) is template, with oligonucleotide 5 '-CCTCCTGCACTGCTGATGCCC-3 ' and oligonucleotide 5 '-CCTTGCTCTTGCTGGCTCTTCC-3 ' is reverse primer, carry out PCR, the PCR condition be 93 ℃ 4 minutes, carried out 35 circulations in 1 minute with 93 ℃ 1 minute, 54 ℃ 1 minute and 72 ℃, last 72 ℃ were extended 5 minutes thereupon.The 436bp purpose fragment that electrophoresis detection obtains.2. probe and mark thereof

With above-mentioned PCR purpose fragment is probe, usefulness α- ³²P-dATP (DuPont company) carries out random primer labelling to it, condition be 37 ℃ 1 hour, MegaprimeDNA Mk system specification sheets (Amersham) is seen in concrete operations.3.cDNA amplified library and trace mark (screening library)

Increased in people's placenta λ gt11cDNA library, clone's number reaches 500,000, good cDNA library trace to the Hybond TM-N+ nylon membrane (Amersham) then will increase, hybridize with blotting membrane after again that mark is the good probe sex change, wash film, put into the sheet folder of band intensifying screen, under-80 ℃, carry out radioautograph, punching after 72 hours with FUJI MEDIAI X-ray film.4. picking clone and primary dcreening operation clone's multiple sieve

Obtain a plurality of primary dcreening operation positive colonies by above-mentioned screening by hybridization, these positive colonies are carried out multiple sieve, finally obtain 16 positive monoclonals with above-mentioned identical probe hybridization and screening process.5. monoclonal evaluation and order-checking

With the mono-clonal phage that obtains in the above-mentioned steps 4 is template, with two positive anti-primers of above-mentioned steps 1 respectively with λ gt11 left and right arms on primer S1 and S2 (S1:5 '-GTTCAACATCAGCCGCTACA-3 '; S2:5 '-CACCAGACCAACTGGTAATG-3 ') amplification of arranging in pairs or groups, carry out agarose gel electrophoresis then, to judge that each clone moves the length of (extension) to 5 ' or 3 ' step from probe, therefrom select to 5 ' and extend the longest and extend the longest clone to 3 ', find that a clone contains the insertion fragment that reaches 2.2Kbp.With this clone's S1-S2 amplified production and pGEM-T ^Carrier (Promega) connects, transformed into escherichia coli JM103, extract plasmid with QIAprepPlasmid test kit (QIAGEN), carry out the mistake of orientations Lieque to inserting fragment, carry out Rapid identification and ordering with PCR to lacking son then with double-stranded nested type disappearance test kit (Pharmacia).Use SwquiTherm EXCEL ^TMDna sequencing kit (Epicentre Technologies) checks order to disappearance of brachymemma successively.Use SequiThermEXCEL ^TMDna sequencing kit (Epicentre Technologies) checks order to disappearance of brachymemma successively, with computer software splicing order, obtain full length cDNA sequence at last, altogether 2069bp, detailed sequence is seen SEQ ID NO:1, and wherein open reading frame is positioned at 139-933 position Nucleotide.

Derive the aminoacid sequence of SOCS-4 according to the cDNA sequence that obtains, totally 264 amino-acid residues, its aminoacid sequence sees SEQ ID NO:2 for details.Expression pattern analysis and the chromosomal localization of embodiment 2 SOCS-4

Multiple Northern (the MNT that organizes ^TM) blotting membrane (16 kinds of tissues) is available from Clontech company, cut the pulsating plasmid vector enzyme of insertion is housed among the embodiment 1 with NotI/SacII, with the disconnected isotopic labeling of carrying out of the 1.6Kbp enzyme section of gained, the marking method of probe is with embodiment 1 step 2, and the user manual operation is pressed in Northern hybridization.

The Northern result of 16 kinds of tissues shows SOCS-4 high expression level in spleen, placenta; In most other tissues, medium as expression amount in thymus gland, prostate gland, ovary, small intestine, colon, heart, lung, liver, skeletal muscle, kidney and the pancreas; The low expression in testis and peripheral blood lymphocyte; And in brain, do not find the expression of CIS-7.

According to international blastn nr database retrieval, with this assignment of genes gene mapping in 3p21.3.

The expression of embodiment 3 SOCS-4 in intestinal bacteria

In this embodiment, be masterplate with people's placenta λ gt11cDNA library, use 5 ' and 3 ' the PCR Oligonucleolide primers of holding to increase the cDNA sequence of coding people SOSC4 corresponding to this dna sequence dna, obtain people SOSC4cDNA as inserting fragment.

5 ' the Oligonucleolide primers sequence of using in the PCR reaction is:

5’-AGATGGATCCATGGGTCCCTCCACTCCTG-3’，

This primer contains the restriction enzyme site of the restricted restriction enzyme of BamHI, is 19 Nucleotide of the people SOSC4 encoding sequence that begun by initiator codon after this restriction enzyme site;

3 ' end primer sequence is:

5’-AAGGAAGCTTTCAGAGCTGGAAGGGGTAC-3’，

This primer contains the part encoding sequence of restriction enzyme site, translation termination and the people SOSC4 of the restricted restriction enzyme of HindIII.

The restriction enzyme site of the restriction enzyme on the primer is corresponding to bacterial expression vector pQE-9 (Qiagen Inc., Chatsworth, CA) restriction enzyme digestion sites on, this plasmid vector coding antibiotics resistance (Ampr), a bacterium replication orgin (ori), an adjustable promotor/operon of IPTG-(P/O), a ribosome bind site (RBS), a 6-histidine mark thing (6-His) and restriction enzyme cloning site.

With BamHI and HindIII digestion pQE-9 carrier and insertion fragment, will insert fragment subsequently and be connected to the pQE-9 carrier and keep open reading frame initial at bacterium RBS.Transform available from Qiagen with connecting mixture subsequently, the E.coli bacterial strain of commodity M15/rep4 by name, M15/rep4 contains the plasmid pREP4 of multiple copied, and it is expressed the lacI repressor and carries kalamycin resistance (Kanr).Screen transformant containing on the LB culture dish of Amp and Kan, the extracting plasmid, the cDNA fragment of sequence verification people SOSC4 has correctly been inserted carrier.

Incubated overnight (O/N) contains the positive transformant clone of required construction in the LB liquid nutrient medium of adding Amp (100 μ g/ml) and Kan (25 μ g/ml).Spend the night (O/N) culture with 1: 100-1: 250 thinning ratio dilution, be inoculated into then in the large volume substratum, culturing cell grows to 600 optical density(OD) (OD600) when the 0.4-0.6, add IPTG (" isopropylthio-") to final concentration be 1mM.By making lacI repressor inactivation, IPTG induces startup P/O to cause gene expression dose to improve.Continued culturing cell 3-4 hour, centrifugal subsequently (6000 * g, 20 minutes).The ultrasonic degradation inclusion body, collecting cell also is dissolved in cell precipitation in the Guanidinium hydrochloride of 6M.After the clarification, by containing under the condition that 6-His marker albumen combines closely making, with nickel-chelate column chromatography purifying dissolved people SOSC4 from solution.With 6M Guanidinium hydrochloride (PH5.0) wash-out people SOSC4 from post.Available several method is the sex change protein precipitation from Guanidinium hydrochloride.Perhaps, use the dialysis step to remove Guanidinium hydrochloride, perhaps isolated purifying protein from nickel-chelate column.Protein behind the purifying is incorporated in second post, has the linear Guanidinium hydrochloride gradient of successively decreasing in this post.Protein denaturation when being attached to this post is used Guanidinium hydrochloride (PH0.5) wash-out subsequently.At last, soluble protein is dialysed with PBS, then protein is kept in the stock solution that final concentration is 10% (w/v) glycerine.

SDS-PAGE glue with 12% carries out electrophoresis, identifies that the molecular weight size of expressing protein is about 29KDa.

In addition, the amino acid that reaches proteic N end and each 10 amino acid length of C end is checked order, find consistent with the protein sequence of inferring with ordinary method.

The expression of embodiment 4 people SOSC4 in eukaryotic cell (Chinese hamster ovary celI strain)

In this embodiment, be masterplate with people's placenta λ gt11cDNA library, use 5 ' and 3 ' the PCR Oligonucleolide primers of holding to increase the cDNA sequence of coding people SOSC4 corresponding to this dna sequence dna, obtain people SOSC4cDNA as inserting fragment.

5 ' the Oligonucleolide primers sequence of using in the PCR reaction is:

5’-AGATAAGCTTATGGGTCCCTCCACTCCTG-3’，

This primer contains the restriction enzyme site of the restricted restriction enzyme of HindIII, is 19 Nucleotide of the people SOSC4 encoding sequence that begun by initiator codon after this restriction enzyme site;

3 ' end primer sequence is:

5’-AAGGGGATCCTCAGAGCTGGAAGGGGTAC-3’，

This primer contains the part encoding sequence of the restriction enzyme site of the restricted restriction enzyme of BamHI, translation termination and people SOSC4.

The restriction enzyme site of the restriction enzyme on the primer is corresponding to the restriction enzyme digestion sites on the expressing cho cell carrier pcDNA3, and this plasmid vector coding antibiotics resistance (Ampr and Neor), a phage replication starting point (fl ori), a virus replication starting point (SV40 ori), T7 promotor, a viral promotors (P-CMV), a Sp6 promotor, a SV40 promotor, a SV40 tailing signal and corresponding polyA order, a BGH tailing signal and a corresponding polyA are in proper order.

With BamHI and HindIII digestion pcDNA3 carrier, will insert fragment subsequently and be connected to the pcDNA3 carrier.Subsequently with connecting mixture Transformed E .coli DH5 α bacterial strain.Screen transformant containing on the LB culture dish of Amp, incubated overnight (O/N) contains the clone of required construction in the LB liquid nutrient medium of adding Amp (100 μ g/ml).The extracting plasmid, and the cDNA fragment of sequence verification people SOSC4 has correctly been inserted carrier.

The plasmid transfection Chinese hamster ovary celI is to adopt lipofection, carries out with Lipofectin (GiBco Life).After the transfection 48 hours, through the lasting G418 pressurization screening in 2-3 week, collecting cell and cell conditioned medium are measured the expressing protein enzyme activity.Remove G418, continuous passage is cultivated; To mixing the clone cell Method of Limited Dilution, select to have the cell subclone of higher protein-active.The above-mentioned positive subclone of a large amount of according to a conventional method cultivations.After 48 hours, beginning collecting cell and supernatant are with ultrasonic degradation method smudge cells.With 50mMTrisHCl (PH7.6) solution that contains 0.05%Triton is balance liquid and elutriant, uses through the Superdex of pre-equilibration G-75 post and collects above-mentioned proteic active peak.Using 50mMTrisHCl (PH8.0) equilibrated DEAE-Sepharose post again, is that elutriant carries out gradient elution with 50mMTrisHCl (PH8.0) solution that contains 0-1M NaCl, collects above-mentioned proteic active peak.Be that dialyzate is dialysed to expressing protein solution with PBS (PH7.4) then.Last freeze-drying is preserved.

SDS-PAGE glue with 12% carries out electrophoresis, identifies that the molecular weight size of expressing protein is 29KDa.

In addition, the amino acid that reaches proteic N end and each 10 amino acid length of C end is checked order, find consistent with the proteic sequence of inferring with ordinary method.

According to above instruction, the present invention can have various improvement and change, and therefore, within the scope of the appended claims, the present invention can implement with concrete described different mode.

Sequence table (1) general information (i) applicant: Fudan University is denomination of invention (ii): cytokine signal transfer arrestin IV is sequence number (ii): 2 (iii) contact addresses:

(A) addressee: Ntd Patent ﹠ Trademark Agency Ltd

(B) street: Building A, No. 27 investment squares, financial street 10 floor

(C) city: Beijing

(D) country: China

(E) postcode: 100032 (vi) the application's data:

(A) application number:

(B) applying date:

(C) classification number: (viii) lawyer/proxy's information:

(A) name: woods Xiao Hong

(B) number of registration: 72002027

(C) reel number: I98597CB (ix) telecommunication information:

(A) phone: 8610-66211834

(B) fax: the information of 8610-66211845 (2) SEQ ID NO:1: (i) sequence signature:

(A) length: 2069bp

(B) type: nucleic acid

(C) chain: strand

( D ) ： ( iii ) ：cDNA ( xi ) ：SEQ ID NO：1 1 GACAAGTGGG GAGCGAGATA CTGGACTAAC TGAGCCCATG GCTGGGTAGA GGGGAAGCTG 61 AGGCTCATAG TGAGATGGGC CAGCCAGAGG TCATGAAACT AGTCTGGGGC ATAGGGGGGT 121 CTCTGGAGAT GGGGAAGCAT GGGTCCCTCC ACTCCTGCTC ATCCACTGCC TTCTAGACCT 181 CGTCCTTTGC TGGCTGTGGA GCGGACTGGG CAGCGGCCCC TGTGGGCCCC GTCCCTGGAA 241 CTGCCCAAGC CAGTCATGCA GCCCTTGCCT GCTGGGGCCT TCCTCGAGGA GGTGGCAGAG 301 GGTACCCCAG CCCAGACAGA GAGTGAGCCA AAGGTGCTGG ACCCAGAGGA GGATCTGCTG 361 TGCATAGCCA AGACCTTCTC CTACCTTCGG GAATCTGGCT GGTATTGGGG TTCCATTACG 421 GCCAGCGAGG CCCGACAACA CCTGCAGAAG ATGCCAGAAG GCACGTTCTT AGTACGTGAC 481 AGCACGCACC CCAGCTACCP GTTCACGCTG TCAGTGAAAA CCACTCGTGG CCCCACCAAT 541 GTACGCATTG AGTATGCCGA CTCCAGCTTC CGTCTGGACT CCAACTGCTT GTCCAGGCCA 601 CGCATCCTGG CCTTTCCGGA TGTGGTCAGC CTTGTGCAGC ACTATGTGGC CTCCTGCACT 661 GCTGATACCC GAAGCGACAG CCCCGATCCT GCTCCCACCC CGGCCCTGCC TATGCCTAAG 721 GAGGATGCGC CTAGTGACCC AGCACTGCCT GCTCCTCCAC CAGCCACTGC TGTACACCTA 781 AAACTGGTGC AGCCCTTTGT ACGCAGAAGC AGTGCCCGCA GCCTGCAACA CCTGTGCCGC 841 CTTGTCATCA ACCGTCTGGT GGCCGACGTG GACTGCCTGC CACTGCCCCG GCGCATGGCC 901 GACTACCTCC GACAGTACCC CTTCCAGCTC TGACTGTACG GGGCAATCTG CCCACCCTCA 961 CCCAGTCGCA CCCTGGAGGG GACATCAGCC CCAGCTGGAC TTGGGCCCCC ACTGTCCCTC1021 CTCCAGGCAT CCTGGTGCCT GCATACCTCT GGCAGCTGGC CCAGGAAGAG CCAGCAAGAG1081 CAAGGCATGG GAGAGGGGAG GTGTCACACA ACTTGGAGGT AAATGCCCCC AGGCCGCATG1141 TGGCTTCATT ATACTGAGCC ATGTGTCAGA GGATGGGGAG ACAGGCAGGA CCTTGTCTCA1201 CCTGTGGGCT GGGCCCAGAC CTCCACTCGC TTGCCTGCCC TGGCCACCTG AACTGTATGG1261 GCACTCTCAG CCCTGGTTTT TCAATCCCCA GGGTCGGGTA GGACCCCTAC TGGCAGCCAG1321 CCTCTGTTTC TGGGAGGATG ACATGCAGAG GAACTGAGAT CGACAGTGAC TAGTGACCCC1381 TTGTTGAGGG GTAAGCCAGG CTAGGGGACT GCACAATTAT ACACTATTTA TTTATTTATT1441 CTCCTTGGGG TTGGTGTCAG GGGCGAGCCA ACCCCACCTC TATGCCCTGA GCCCTGGTAG1501 TCCAGAGACC CCAACTCTGC CCTGGCTTCT CTGGTTCTTC CCTGTGGAAA GCCCATCCTG1561 AGACATCTTG CTGGAACCAA GGCAATCCTG GATGTCCTGG TACTGACCCA CCCGTCTGTG1621 AATGTGTCCA CTCTCTTCTG CCCCCAGCCA TATTTGGGGA GGATGGACAA CTACAATAGG1681 TAAGAAAATG CAGCCGGAGC CTCAGTCCCC AGCAGAGCCT GTGTCTCACC CCCTCACAGG1741 ACAGAGCTGT ATCTGCATAG AGCTGGTCTC ACTGTGGCGC AGGCCCCGGG GGGAGTGCCT1801 GTGCTGTCAG GAAGAGGGGG TGCTGGTTTG AGGGCCACCA CTGCAGTTCT GCTAGGTCTG1861 CTTCCTGCCC AGGAAGGTGC CTGCACATGA GAGGAGAGAA ATACACGTCT GATAAGACTT1921 CATGAAATAA TAATTATAGC AAAGAACAGT TTGGTGGTCT TTTCTCTTCC ACTGATTTTT1981 CTGTAATGAC ATTATACCTT TATTACCTCT TTATTTTATT ACCTCTATAA TAAAATGATA2041 CCTTTCATGT AAAAAAAAAA AAAAAAAAA ( 2 ) SEQ ID NO：2：

(i) sequence signature:

(A) length: 264 amino acid

(B) type: amino acid

(C) topological framework: linearity

(iii) molecule-type: polypeptide

(xi) sequence description: SEQ ID NO:2 1 MGPSTPAHPLPSRPRPLLAVERTGQRPLWAPSLELPKPVMQPLPAGAFLEEVAEGT PAQT 61 ESEPKVLDPEEDLLCIAKTFSYLRESGWYWGSITASEARQHLQKMPEGTFLVRDST HPSY121 LFTLSVKTTRGPTNVRIEYADSSFRLDSNCLSRPRILAFPDVVSLVQHYVASCTAD TRSD181 SPDPAPTPALPMPKEDAPSDPALPAPPPATAVHLKLVQPFVRRSSARSLQHLCRLV INRL241 VADVDCLPLPRRMADYLRQYPFQL

Claims (18)

1, a kind of isolated polynucleotide, its coding has the polypeptide of deduced amino acid of SEQ ID NO:2 or fragment, analogue or the derivative of described polypeptide, and described fragment, analogue or derivative have the biological function identical with full-length polypeptide.

2, polynucleotide as claimed in claim 1, wherein these polynucleotide are DNA.

3, polynucleotide as claimed in claim 1, wherein these polynucleotide are RNA.

4, polynucleotide as claimed in claim 1, wherein these polynucleotide are genomic dnas.

5, polynucleotide as claimed in claim 1, its coding have the polypeptide of the deduced amino acid of SEQ ID NO:2.

6, polynucleotide as claimed in claim 1, it has the encoding sequence shown in the SEQ ID NO:1.

7, the carrier that contains the described polynucleotide of claim 2.

8, a kind of host cell, it uses carrier conversion, transfection or the transduction of claim 7.

9, host cell as claimed in claim 8, this host cell are prokaryotic cell prokaryocyte.

10, host cell as claimed in claim 9, this host cell are intestinal bacteria.

11, host cell as claimed in claim 8, this host cell are eukaryotic cell.

12, host cell as claimed in claim 11, this host cell are COS-7, CHO, Hela cell.

13, host cell as claimed in claim 11, this host cell are yeast cell.

14, produce the method for polypeptide, be included in the host cell of cultivating claim 8-13 under the appropriate condition, make it to express described polypeptide.

15, a peptide species, described polypeptide are to have the polypeptide of deduced amino acid of SEQ ID NO:2 or fragment, analogue or the derivative of described polypeptide, and described fragment, analogue or derivative have the biological function identical with full-length polypeptide.

16, polypeptide as claimed in claim 15, described polypeptide has the deduced amino acid of SEQ ID NO:2.

17, the antibody of anti-claim 15 or 16 described polypeptide.

18, claim 15 or the 16 described polypeptide application in the medicine of preparation diagnosis and treatment and its relative disease.