CN104447964A - Vaccine medicine target SIM resisting Kaposi's sarcoma-associated herpesvirus - Google Patents

Abstract

The invention belongs to the field of biological medicines, relates to a vaccine medicine target SIM (SUMO-Interacting Motif) resisting Kaposi's sarcoma-associated herpesvirus (KSHV), and concretely relates to an SIM polypeptide sequence resisting Kaposi's sarcoma-associated herpesvirus and application thereof to prepare vaccines. The SIM domain has important effect on maintaining incubation-period infection such as stable and continuous heredity and the like of genome of Kaposi's sarcoma-associated herpesvirus. The SIM domain is capable of recognizing transcription silencing complex of multiple transcription factors such as SUMO-2 modified KAP1 and Sin3A and the like. Deletion of the SIM domain influences stable heredity and potential infection of KSHV virus genome, and improves the efficiency of KSHV virus starting the cracking life cycle and destroying host cells under a microenvironment stress condition. The SIM domain can be used as a treatment target for resisting Kaposi's sarcoma-associated herpesvirus. The invention also relates to an artificial reconstructed nucleotide sequence of a start epitope of the polypeptide sequence, nucleotide sequences corresponding to different assembling sequences of the whole SIM subfunctional domain and encoded protein products thereof, and potential application of the polypeptide sequence to treatment resisting Kaposi's sarcoma-associated herpesvirus.

Description

The pharmaceutical vaccine target spot SIM of anti-card Perth sarcoma herpes virus

Technical field

The invention belongs to field of biomedicine technology, relate to the development & application of gene, the SIM peptide sequence being specifically related to a kind of anti-Perth sarcoma herpes virus is as the purposes of the immunotherapy of vaccine and drug target.

Background technology

Prior art discloses the main pathogens that card Perth sarcoma herpes virus KSHV is multiple human malignancies, is the main cause of one of the most common 4 kinds of infectious tumours.Research display, KSHV infect the tumor invasion such as AIDS patient relevant to HIV, organ transplantation immunosuppressed patient and the low the elderly of partial immunity and lethality rate closely related.In view of HIV rate rise and organ transplantation/autoimmune patient's immunotherapeutic day by day extensive, intervention KSHV Pathogenic pathway become a kind of in the urgent need to reply behave.In global range, in countries of sub-Saharan Africa, Prevalent district KSHV detects up to 80%, and the U.S. reaches 20%.According to estimates, the acquired immune deficiency syndrome (AIDS) death of about 14% is caused by the concurrent cancer of the most common card Perth sarcoma (4%) relevant to KSHV.According to another statistics, the whole world every year about 60,000 examples lymphoma primary effusion (PEL) diagnosis in, aids related lymphoma accounts for 3-4%.The more important thing is, in association area, not yet set up the methods for the treatment of or the vaccine that cause malignant tumour for KSHV at present.Research display, because KSHV can utilize a few virogene viral genome be attached to the karyomit(e) of host and escape host immune system of supervision and set up long-term latent infection, therefore, once infected, patient will carry virus all the life.

Similar to other simplexvirus, KSHV virus has latent period and two life cycles of burst times.In the latent infection phase, KSHV virus only has a small amount of several viral gene expression, which includes the relevant nuclear antigen LANA that hides.There are some researches prove that LANA is the key protein being responsible for safeguarding that KSHV virus lays dormant infects.Research show LANA be one containing 1162 amino acid whose multi-functional viral proteins, it is not only virus genomic with KSHV stablely continues heredity and sets up latent infection closely related, and has direct effect with degraded tumor suppressor protein such as p53 and VHL etc.But, so far, not yet have the therapeutic vaccine designed for KSHV virus infection characteristic.Therefore, the therapeutic vaccine infected for KSHV virus lays dormant is researched and developed most important.

Summary of the invention

The object of the invention is the defect for overcoming prior art, the pharmaceutical vaccine target spot SIM of anti-card Perth sarcoma herpes virus is provided, be specifically related to by the card Perth sarcoma herpes virus (KSHV) relevant to acquired immune deficiency syndrome (AIDS) encode containing SUMO-2 specific combination functional domain (SIM suMO- interacting motif) peptide sequence.

A further object of the present invention is, provides the SIM peptide sequence of described anti-Perth sarcoma herpes virus and the purposes in preparation immunotherapy vaccine and drug target thereof.

The present invention is based on a large amount of early-stage Study to find, the peculiar SIM functional domain of the relevant nuclear antigen LANA that hides.This SIM structural domain is that KSHV encoding viral to be correlated with the functional domain of nuclear antigen LANA latent period, stable to the genome of card Perth sarcoma herpes virus, continue heredity etc. and safeguard that latent period infects and play an important role.This peptide sequence can identify the Transcriptional Silencing mixture comprising and modified the multiple transcription factors such as KAP1 and Sin3A by SUMO-2.

The present invention our experiments show that, aforementioned polypeptides sequence deletion can affect the virus genomic genetic stability of KSHV and latent infection, and closing of SIM site can promote card Perth sarcoma virus promoter lysing cell life cycle and the efficiency destroying host cell.

Further experiment of the present invention shows that blocking-up latent infection not only can stop infection subsequently, and can reduce the progress of tumour, and this SIM structural domain can be used as the candidate therapeutic target spot of anti-card Perth sarcoma herpes virus.

The functional domain SIM be combined with SUMO-2 of the present invention is a critical sites of LANA albumen, and it comprises the full-length polypeptide of 61 amino-acid residues shown in coding SEQID No:1;

1) this sequence cDNA sheet segment length is 183bp, 61 amino acid of encoding, and molecular weight is the albumen of 9kDa;

2) containing the sub-functional domain that 4 SUMO-2 combine, there is 1 IYV, 1 ISI and 2 SSS.

The invention still further relates to the protein of the artificial reconstructed nucleotide sequence of the epi-position of setting out of described peptide sequence, the different assembling sequence corresponding nucleotide sequence of the sub-functional domain of whole SIM and its coding, and the purposes in described gene and the treatment preparation of protein in the anti-card Perth sarcoma herpes virus of preparation.

As in an experiment prove, SIM polypeptide protein of the present invention can with a large amount of Transcriptional Silencings identify, the disappearance in this site can destroy the ability that LANA safeguards viral genome genetic stability effectively.

In addition, the LANA mutant loss of SIM site deletion of the present invention checks burst times modulin RTA gene transcript expression, and can in cell effectively cracking destroy tumour cell.

SIM polypeptide vaccine drug target of the present invention has high specific, the validity that anti-KSHV virus lays dormant infects, and institute produces antibody or small-molecule drug and resists KSHV virus infection and have very high potential suppression efficiency.

SIM polypeptide protein of the present invention can be used as antigen and produces neutralizing antibody and as the small-molecule chemical medicine obtained in conjunction with Sites Screening, further for the preparation of the immunological reagent of prevention and therapy KSHV virus associated-diseases, as, the vaccine of anti-card Perth sarcoma virus infection in latent period.

For the ease of understanding, by by concrete drawings and Examples, the present invention is described in detail below.It needs to be noted, specific examples and accompanying drawing are only to illustrate, obvious those of ordinary skill in the art according to illustrating, can make various correction and change to the present invention herein within the scope of the invention, and these are revised and change and also include in scope of the present invention.

Accompanying drawing illustrates:

Fig. 1 illustrates in card Perth sarcoma herpes virus coding LANA containing 61 amino acid polypeptide sequence (i.e. SEQID NO:1) in SIM site and 183 base sequences (i.e. SEQ ID NO:2) corresponding to encoding histone thereof.

Fig. 2 illustrates containing the distribution situation in 61 aminoacid sequences (i.e. SEQ ID NO:1) Zhong Liangge Dare SIM (SIM1 and SIM2) of SIM site polypeptide and neighbouring acidic amino acid section (AD) space structure.

Fig. 3 is the Intracellular transcription factor coniplexes with SIM structural domain specific combination,

Upper figure: SDS-PAGE detects the coomassie brilliant blue staining result of prokaryotic expression protein GST and GST-SIM in conjunction with albumen composition in 293 cells; Figure below: with the mass spectrum sequencing result of SIM structural domain specific combination b1, b2, b3 and b4 albumen in upper figure.

Fig. 4 is that SIM disappearance reduces LANA and KAP1 and Sin3A protein binding capacity,

Wherein, prokaryotic expression protein GST and GST merges the difference of wild-type (WT) or SIM disappearance (Δ SIM) LANA (figure below is coomassie brilliant blue staining result) and KAP1, Sin3A or DNA-PKc binding ability (upper figure is immunoblot results).

Fig. 5 is that SIM disappearance reduces the stable characteristic that goes down to posterity of LANA mediation TR,

Wherein,

A: karyomit(e) co-precipitation detects SIM site and acid section AD lacks the impact of LANA in conjunction with TR ability; B: the process of purinase element is containing Different L ANA mutant and TR-Puro ^rthe cell colonies analytical results of 293 cells after 14 days.

Fig. 6 shows SIM disappearance and causes LANA to regulate the forfeiture of key protein RTA genetic transcription rejection ability to burst times, and under the normal oxygen supply of fluorescent reporter gene analysis and hypoxia condition, Different L ANA mutant is on the impact of Rta gene promoter transcription ability.

Fig. 7 shows SIM deletion mutant dominant negative effect and activates KSHV viral dna replication,

Wherein, containing the Different L ANA mutant of SIM disappearance to the activation of 293 cells of carrying wild-type KSHV virus.

Embodiment

With the determination of SUMO-2 specific combination SIM structural domain and sequential analysis in embodiment 1:LANA

1. with the determination in the LANA SIM site of SUMO-2 specific combination

Select the different section LANA mutant of band myc label and SUMO-1 or SUMO-2 coexpression in human embryo kidney 293 cells of band FLAG label, then co-immunoprecipitation determines that special high-level binding site is the 233 to 340 stretches of amino acids in LANA with SUMO-2.Adopt disappearance and point mutation process to be defined as Liang Ge Dare SIM (i.e. 4 little sub-SIM) binding domains, be respectively ₂₄₄-IYVGSSS- ₂₅₀with ₂₆₄-ISIGSSS- ₂₇₀(as shown in Fig. 1 SEQ ID NO:1), corresponding nucleotide sequence (as shown in Fig. 1 SEQ ID NO:2).

2. containing space binding domain and the sequential analysis of SIM site functional polypeptide

Space binding domain and sequence alignment analysis is carried out for LANA the 233 to 340 aminoacid sequence containing SIM site, result shows its 240 to 300 amino acid (SEQ ID NO:2) and forms relatively independent " saddle type " space structure territory, near it, the acidic amino acid section of 86%QDE content is " encircling type " contiguous SIM structural domain, as shown in Figure 2.

The separation and purification of embodiment 2:SIM polypeptide protein and the qualification of ICBP mixture thereof

1.GST merges the structure of SIM polypeptide protein genetic engineering bacterium and the expression and purification of its proteins encoded

By the pcr amplification product containing SIM peptide sequence (240-300 amino acid) after Bgl II and EcoRI enzyme cut, be connected to coli expression carrier pGEX-2TK, competence transformation of E. coli BL21 strain, be after the Screening of Media of 100 μ g/ml amicillin resistances at final concentration, obtain containing recombinant plasmid GST-SIM prokaryotic expression engineering bacteria GST-SIM/BL21.This recombinant strain, in glycerol stocks mode, backs up (Beijing) in Chinese Academy of Sciences's Organism Depositary preservation.This recombinant strain induces 6 hours through 30 DEG C of IPTG, obtains the fusion recombinant protein (with GST protein fusion) of GST-SIM, through the gsh affinity purification fusion rotein (the upper figure of Fig. 3) of Specific adsorption.

Concrete steps are as follows:

1) recombinant protein preparation of samples:

A) contain the colibacillus engineering of expressing target protein at 5ml LB inoculation of medium, 37 DEG C of activation are spent the night.

B) 1/100 inoculum size is transferred to fresh LB37 DEG C and cultivates to close to logarithmic phase, and adding final concentration is that 1mM IPTG was 30 DEG C of inductions 6 hours.

C) bacterium liquid 4 DEG C is collected, 5000rpm × 5min.

D) add the resuspended bacterium liquid of 1 × STE of the precooling of certain volume, 4 DEG C, 5000rpm × 5min, removes supernatant.

E) add NETN damping fluid and the resuspended bacterium liquid of 75 μ l1M DTT/1ml STE that 1.5ml contains proteinase inhibitor, carry out ultrasonic wave (40% intensity, 10 seconds, 10 seconds, interval, 50 times) on ice.

F) 4 DEG C, 10,000rpm × 10min, supernatant proceeds to a new 50ml centrifuge tube, adds STE and the affine pearl of 100 μ l gsh that 1.5ml contains 10%Triton-X100, and 4 DEG C rotate 2hr.

G) sample is transferred to 2ml centrifuge tube, collects 4 DEG C, pearl, 3000rpm × 5min.

H) the NETN damping fluid containing proteinase inhibitor washes 5 times, each 1ml.

I) NETN damping fluid is resuspended, puts 4 DEG C and saves backup.

2) reagent prepares:

STE damping fluid: 100mM NaCl, 10mM Tris, 1mM EDTA, pH=7.5

NETN damping fluid: 0.5%NP40,20mM Tris pH=8.0,1mM EDTA, 100mM NaCl

RIPA damping fluid: 1%NP40,10mM Tris pH=7.5,2mM EDTA, 150mM NaCl

2. with the mass spectrum sequencing analysis of SIM polypeptide in conjunction with intracellular protein

10 ⁸after 4 DEG C, GST and the GST-SIM albumen that individual human embryo kidney (HEK) 293 nuclear RIPA lysate (NE) and the above-mentioned purifying of 100 μ l obtain rotates overnight incubation, wash 5 times, each 4 DEG C, 3000rpm × 5min with 1ml containing proteinase inhibitor TBS damping fluid.Then the 100 DEG C of 5min sex change of SDS sample-loading buffer are added, 4%-15% gradient glue concentration SDS-PAGE electrophoretic separation (shown in figure as upper in Fig. 3); By distinctive obvious 4 protein band b1 in the nucleus lysate that is combined with GST-SIM, b2, b3 and b4 cut recovery, and after being transferred to centrifuge tube, 5% Glacial acetic acid damping fluid send MALDI-TOF mass spectroscopy; Result shows, in b1 band, high-abundance proteins is DNA-PKc; In b2 band, high-abundance proteins is CBP and p300; In b3 band, high-abundance proteins is Sin3A; In b4 band, high-abundance proteins is KAP1 (as shown in Fig. 3 figure below);

3.KAP1 and Sin3A is that SIM polypeptide is in conjunction with the key protein in mixture

For determine mass spectroscopy in 5 kinds of albumen during SIM polypeptide is in conjunction with mixture which or several are key proteins, as step 1 purifying obtain GST, GST-LANA1-329WT and GST-LANA1-329 Δ SIM fusion rotein respectively with 10 ⁶after individual human embryo kidney 293 cells RIPA lysate 4 DEG C rotates overnight incubation, wash 5 times, each 4 DEG C, 3000rpm × 5min with 1ml containing proteinase inhibitor TBS damping fluid; Then the 100 DEG C of 5min sex change of SDS sample-loading buffer are added, after 6% gradient glue concentration SDS-PAGE electrophoretic separation, the protein immunoblot carrying out anti-DNA-PKc, CBP, p300, Sin3A and KAP1 respectively detects, result shows, KAP1 and Sin3A is that SIM polypeptide is in conjunction with the key protein (shown in figure as upper in Fig. 4) in mixture.

GST co-immunoprecipitation experiment step:

1) regulating total protein concentration to be 1mg volume with RIPA cell pyrolysis liquid is 1ml, GST or GST adding 50 μ l merges egg

White suspension, 4 DEG C of slow rolling 1-3 hour;

2) 4 DEG C, centrifugal 5 minutes of 3,000rpm, removes supernatant;

3) add 1ml washing lotion NETN, resuspended albumen pearl, 4 DEG C are slowly rolled 20 minutes;

4) above-mentioned two step five times are repeated;

5) collect albumen pearl, remove the washing lotion of tracer level as far as possible;

6) sample-loading buffer of 25 μ l is added; 100 DEG C of sex change 3 minutes,

7) 12,000g centrifugal 20 seconds;

8) supernatant carries out SDS-polyacrylamide gel electrophoresis;

9) pvdf membrane soaks 1min in methyl alcohol, puts into transferring film damping fluid infiltrate to transferring film with two layers of filter paper, sponge pad;

10) sponge, filter paper, gel, pvdf membrane, filter paper, sponge follow procedure are stacked, 100v transferring film 60min;

11) 5% skim-milk (PBS preparation) is closed, room temperature, 30min;

12) TBST washs three times, 5min/ time, with one-level rabbit source antibody (1:5000, PBS dilute, Huaan, Hangzhou), and 4 DEG C of overnight incubation;

13) TBST washs three times, 5min/ time, with the goat anti-rabbit antibody incubated at room 1h of HRP mark; (1:5000, PBS dilute, Ai Bimate) TBST washs three times, 5min/ time;

14) ECL test kit development.

Embodiment 3:SIM disappearance causes LANA to safeguard the forfeiture of KSHV viral genome genetic stability function

1. karyomit(e) co-immunoprecipitation detects SIM site deletion to the impact of LANA DNA binding ability

For SIM site determined in embodiment 1 and acidic amino acid section AD, after building the LANA mutant of SIM site and/or AD section or the only band myc label of C-terminal, respectively with recombinant plasmid pBS-Puro-TR cotransfection 293 cell carrying KSHV viral genome TR (terminal repeat) and purinase element resistance, after 48 hours, as follows the nuclear extract of transfectional cell is carried out the analysis of karyomit(e) co-immunoprecipitation respectively, result as shown in Figure 5A, shows that the DNA binding ability of SIM site deletion to LANA does not make significant difference.

The nuclear extract of transfectional cell is carried out the analysis of karyomit(e) co-immunoprecipitation respectively:

10xHEPES-NaOH damping fluid:

1M Hepes pH=7.8

11% methanol solution (approx)

Time crosslinked, working fluid concentration is 1X

This solution is 10X mother liquor, matching while using during experiment, and working fluid concentration is 1X;

The molten Cell Buffer of ChIP:

50mM Tris-Cl pH8.0

10mM EDTA

1％SDS

+ proteinase inhibitor;

RIPA damping fluid (1X):

10mM Tris-Cl pH8.0

1mM EDTA

0.5mM EGTA

140mM NaCl

1％Triton X100

0.1% Sodium desoxycholate

0.1％SDS

+ proteinase inhibitor;

Digesting damping fluid:

mM Tris-Cl pH8.0

1mM EDTA

100mM NaCl

0.5％SDS

(adding 100ug/mL K proteolytic enzyme).

Experimental procedure:

1) by 2x10 ⁷transfected cells adds the HEPES/ formaldehyde solution of newly joining, and is that 1% formaldehyde (adds the HEPES-NaOH-formaldehyde solution of 0.5ml 11% to final concentration; Matching while using on pretreatment),

2) incubated at room 10 minutes,

3) after peptic cell, 4 DEG C, centrifugal 5 minutes of 2000rpm,

4) PBS buffer solution material, centrifugal, repeats twice,

5) add the CHIP lysis buffer (the every 10mg total protein of 1mL) containing proteinase inhibitor, sample divided and is filled in two eppendorf centrifuge tubes, each about 500uL,

6) sample is carried out ultrasonic disruption, to obtain the chromatin that length is about 500-1200bp, ultrasonic disruption process is all carried out on ice (referral procedure: ultrasonication 2 seconds, suspends 10 seconds, 6 circulations),

7) high speed centrifugation ultrasonic disruption product, merges the product in two centrifuge tubes of each sample,

8) get 5uL sample, electrophoresis detection size in 1% agarose gel, if stripe size is about 3kb, then carry out step (band usually not separating crosslinked chromatin electrophoresis is greater than 3kb) below,

9) get a part of sample to contrast (5-20%) as the input of PCR,,,,

10) prepurification chromatin: use resin, as Protein A, immunocomplex is obtained after immunoprecipitation, often pipe 40uL packing pre-wash Protein A agar column and add 0.5X RIPA damping fluid+proteinase inhibitor dilute sample, according to electrophoresis result, add about 100-200uL ultrasonic disruption product, prepurification mixture cumulative volume is 1ml, 4 DEG C, upset shaking table hatches more than 1 hour, removes the non-specific impurity that can be combined on resin; Preferably a sample average is divided to multitube prepurification in the present invention, with obtain by after a sample prepurification again packing to the better effect of multitube; Of short duration centrifugal (2000rpm, 1min), supernatant is transferred in new centrifuge tube, adds suitable antibody (about 3ug, depends on antibody concentration), 4 DEG C, and upset shaking table is hatched 3 hours or spends the night;

11) the DNA-albumen composition under immunoprecipitation is added the agar column that new pre-wash is crossed, hatch 1.5 hours for 4 DEG C;

12) of short duration centrifugal (2000rpm, 1min) mixture, by 800ul, 0.5X RIPA buffer solution precipitation, repeats 3 times, (add damping fluid, the mixing of vibration centrifuge tube, of short duration centrifugal, remove supernatant, repeated washing);

13) add 300ul digesting damping fluid, 65 DEG C hatch 4 hours-spend the night, remove crosslinked;

14) often pipe adds equal-volume phenol/chloroform/isoamylalchohol, vortex oscillation, and 4 DEG C of high speed centrifugations 10 minutes, shift liquid supernatant to new centrifuge tube, alcohol settling DNA, because the DNA amount obtained is little, adopts glycogen or pellet paint as carrier in the present invention;

15) 4 DEG C, most high speed centrifugation 30 minutes, removes supernatant, drying precipitated, by the resuspended precipitation of 20ul water, by this template as PCR, and each reaction 2ul.

2. cell colonies experimental analysis SIM site deletion mediates the impact of KSHV viral genome genetic stability on LANA

The cell colonies analysis that purinase element carries out genetic stability is added with the transfectional cell of gained in embodiment 3, after process in 14 days, result shows, and significantly reduces the ability (as shown in Figure 5 B) that LANA mediates KSHV viral genome genetic stability after SIM site deletion.

Embodiment 4SIM site is infected the LANA suppression KSHV virolysis phase and is played a crucial role

1. luciferase reporter gene analyzing and testing SIM site deletion suppresses the impact of burst times activator RTA genetic transcription ability on LANA

To adopt in embodiment 3 similarity method will to express different SIM site and/or AD section and the LANA mutant of band myc label and burst times activator RTA promoters driven luciferase reporter gene pRta-luc cotransfection 293 cell, transfection carries out normal oxygen and low oxygen concentration process to transfectional cell after 24 hours respectively, after process in 24 hours, lysis buffer (Catalog#K801-200Promega company) cell is reported respectively with 200 μ l, 40 μ l lysis supernatants and 25 μ l fluorescence assay substrate reactions, with OpticompI Luminometer (MGM Instruments, Inc.) fluorescence intensity, active in contrast internal reference with the 450nm light absorption value of beta-galactosidase enzymes degraded ONPG substrate, using the multiple relative to Reporter gene vector itself as relative fluorescence (RLU), acquired results is that three experiments repeat, result shows, and under normal oxygen supply condition, significantly reduces LANA mediate cytolysis phase activator RTA Transcription inhibition ability, and have no significant difference (as shown in Figure 6) under anoxia condition after SIM site deletion.

The LANA mutant dominant negative negative regulator of 2.SIM site deletion activates the genome duplication of KSHV virus

To adopt in embodiment 3 similarity method by the SIM site of various dose and/or AD section and the LANA mutant expression plasmid transfection of band myc label to 293 stable cell lines (293/Bac36) carrying KSHV virus, transfection is after 48 hours, use lysis buffer [10mM Tris-HCl pH8.0 respectively, 150mM NaCl, 10mM EDTA, 1%SDS] and Proteinase K to transfectional cell carry out cracking extract virus genom DNA.Gained viral DNA is with specific gene primer K8:5 '-CAAGCTCGCTGTTGTCAACC-3 '; 5 '-GTCCTCTTGGTGGTGTGTGA-3 '; TR:5 '-GGGGCGCGGGGTGTTCACGTAGT-3 '; 5 '-GGGGGCGCCCTCTCTCTACT-3 ' carries out quantitative PCR, with GAPDH:5 '-ACGACCACTTTGTCAAGCTC-3 '; 5 '-GGTCTACATGGCAACTGTGA-3 ' is internal reference; Result shows, and SIM site deletion improves the virus genomic copy number of KSHV significantly, and AD segment deletion can promote that the virus genomic duplicate copy number of KSHV that SIM site deletion causes rises (as shown in Figure 7) further.

Claims (7)

1. a peptide sequence for anti-card Perth sarcoma herpes virus, it is characterized in that, its aminoacid sequence is as shown in SEQID No:1;

1) this sequence cDNA sheet segment length is 183bp, 61 amino acid of encoding, and molecular weight is the albumen of 9kDa;

2) containing the sub-functional domain that 4 SUMO-2 combine, there is 1 IYV, 1 ISI and 2 SSS.

2. a nucleic acid, is characterized in that, this nucleic acid sequence encoding protein according to claim 1.

3. by the peptide sequence of anti-card Perth sarcoma herpes virus according to claim 1, it is characterized in that, it also comprises: have the polypeptide of the aminoacid sequence of SEQID No:1 or its artificial protein or the nucleotide sequence needed for corresponding encoded.

4. the peptide sequence described in claim 1,2 or 3 is preparing the purposes in the genomic stability preparation of maintaining card Perth sarcoma virus.

5. the purposes of the peptide sequence described in claim 1,2 or 3 in the vaccine of preparation anti-card Perth sarcoma virus infection in latent period.

6. by purposes according to claim 5, wherein, containing the closed promotion card Perth sarcoma virus promoter lysing cell life cycle in SUMO-2 specific combination functional domain (SIM) site.

7. the peptide sequence described in claim 1,2 or 3 obtains the purposes in the small-molecule chemical medicine of anti-card Perth sarcoma virus infection in latent period in screening, and wherein said peptide sequence is as drug target.