Antigen epitopes of animal coronaviruses:a mini-review

Coronaviruses are widespread in nature and can infect mammals and poultry,making them a public health concern.Globally,prevention and control of emerging and re-emerging animal coronaviruses is a great challenge.The mecha-nisms of virus-mediated immune responses have important implications for research on virus prevention and control.The antigenic epitope is a chemical group capable of stimulating the production of antibodies or sensitized lympho-cytes,playing an important role in antiviral immune responses.Thus,it can shed light on the development of diagnos-tic methods and novel vaccines.Here,we have reviewed advances in animal coronavirus antigenic epitope research,aiming to provide a reference for the prevention and control of animal and human coronaviruses.

Recent advances in the study of epitopes,allergens and immunologic cross-reactivity of edible mango

Mango(Mangifera indica L.)is a tropical fruit that is widely consumed as both fresh fruits and processed products around the world.The high incidence of mango allergy,on the other hand,has sparked widespread concern.Therefore,a summary and analysis of the current status and issues in mango allergen research can guide in-depth study on the mechanism of mango allergy and reveal effective desensitization methods.We described the incidence of fruit allergy,as well as the mechanism and clinical symptoms of mango allergy,in this review.We also looked into the structural properties of mango allergens,the effect of processing methods on mango allergens,prediction methods for mango allergen epitopes,and the current state of research on mango cross-reactive allergens and preventive measures.Finally,the research directions and ideas for the future are proposed and discussed.

Identification and epitope mapping of anti-p72 single-chain antibody against African swine fever virus based on phage display antibody library

African swine fever virus(ASFV)is a lethal pathogen that causes severe threats to the global swine industry and it has already had catastrophic socio-economic effects.To date,no licensed prophylactic vaccine exists.Limited knowledge exists about the major immunogens of ASFV and the epitope mapping of the key antigens.As such,there is a considerable requirement to understand the functional monoclonal antibodies(mAbs)and the epitope mapping may be of utmost importance in our understanding of immune responses and designing improved vaccines,therapeutics,and diagnostics.In this study,we generated an ASFV antibody phage-display library from ASFV convalescent swine PBMCs,further screened a specific ASFV major capsid protein(p72)single-chain antibody and fused with an IgG Fc fragment(scFv-83-Fc),which is a specific recognition antibody against ASFV Pig/HLJ/2018 strain.Using the scFv-83-Fc mAb,we selected a conserved epitope peptide(221MTGYKH226)of p72 retrieved from a phage-displayed random peptide library.Moreover,flow cytometry and cell uptake experiments demonstrated that the epitope peptide can significantly promote BMDCs maturation in vitro and could be effectively uptaken by DCs,which indicated its potential application in vaccine and diagnostic reagent development.Overall,this study provided a valuable platform for identifying targets for ASFV vaccine development,as well as to facilitate the optimization design of subunit vaccine and diagnostic reagents.

The amino acids differences in epitopes may promote the different allergenicity of ovomucoid derived from hen eggs and quail eggs

Quail egg ovomucoid can inhibit activation of basophils and eosinophils,while hen egg ovomucoid has been shown to be a major allergen,named Gal d 1.At present,the differences in structure and function between two ovomucoid are unclear.We found the homology of ovomucoid in quail eggs and hen eggs reached77%.Compared with hen egg ovomucoid,the distribution of secondary structure was different in AA52-53,AA57-58,AA66-68,AA71-72,AA131-133,AA139-140,AA157-159 and AA184-185.Among 9 epitopes of egg ovomucoid,there were different amino acids from quail egg ovomucoid in 8 epitopes.Recombination quail egg ovomucoid had trypsin inhibition activity and quail egg ovomucoid didn't specifically bind to serum of eggs allergic patients.Quail egg ovomucoid can significantly inhibit RBL-2 H3 cells degranulation and protect cells morphology to a certain extent,indicating quail egg ovomucoid can inhibit cells activation and have potential anti-allergic effects,which is related to trypsin inhibitory activity.The difference in sensitization compare to hen egg ovomucoid may be due to amino acids differences affecting protein structure by changing antigenic epitopes.

Rapid, accurate and serotype independent pipeline for in silico epitope mapping of SARS-CoV-2 antigens: a combined machine learning and Chou’s pseudo amino acid composition method

Here,a new integrated machine learning and Chou’s pseudo amino acid composition method has been proposed for in silico epitope mapping of severe acute respiratorysyndrome-like coronavirus antigens.For this,a training dataset including 266 linear B-cell epitopes,1,267 T-cell epitopes and 1,280 non-epitopes were prepared.The epitope sequences were then converted to numerical vectors using Chou’s pseudo amino acid composition method.The vectors were then introduced to the support vector machine,random forest,artificial neural network,and K-nearest neighbor algorithms for the classification process.The algorithm with the highest performance was selected for the epitope mapping procedure.Based on the obtained results,the random forest algorithm was the most accurate classifier with an accuracy of 0.934 followed by K-nearest neighbor,artificial neural network,and support vector machine respectively.Furthermore,the efficacies of predicted epitopes by the trained random forest algorithm were assessed through their antigenicity potential as well as affinity to human B cell receptor and MHC-I/II alleles using the VaxiJen score and molecular docking,respectively.It was also clear that the predicted epitopes especially the B-cell epitopes had high antigenicity potentials and good affinities to the protein targets.According to the results,the suggested method can be considered for developing specific epitope predictor software as well as an accelerator pipeline for designing serotype independent vaccine against the virus.

Prediction of promiscuous T cell epitopes in RNA dependent RNA polymerase of Chikungunya virus

Objective: To explore RNA dependent RNA polymerase of Chikungunya virus(CHIKV) and develop T cell based epitopes with high antigenicity and good binding affinity for the human leukocyte antigen(HLA) classes as targets for epitopes based CHIKV vaccine. Methods: In this study we downloaded 371 non-structural protein 4 protein sequences of CHIKV belonging to different regions of the world from the US National Institute of Allergy and Infectious Diseases(NIAID) virus pathogen resource database. All the sequences were aligned by using CLUSTALW software and a consensus sequence was developed by using Uni Pro U Gene Software version 1.2.1. PropredⅠand Propred software were used to predict HLAⅠ and HLAⅡ binding promiscuous epitopes from the consensus sequence of non-structural protein 4 protein. The predicted epitopes were analyzed to determine their antigenicity through Vaxijen server version 2.0. All the HLAⅠ binding epitopes were scanned to determine their immunogenic potential through the Immune Epitope Database(IEDB). All the predicted epitopes of our study were fed to IEDB database to determine whether they had been tested earlier. Results: Twenty two HLA class Ⅱ epitopes and eight HLA classⅠepitopes were predicted. The promiscuous epitopes WMNMEVKII at position 486–494 and VRRLNAVLL at 331–339 were found to bind with 37 and 36 of the 51 HLA class Ⅱ alleles respectively. Epitope MANRSRYQS at position 58–66 and epitopes YQSRKVENM at positions 64–72 were predicted to bind with 12 and 9 HLAⅠI alleles with antigenicity scores of 0.754 9 and 1.013 0 respectively. Epitope YSPPINVRL was predicted to bind 18 HLAⅠ alleles and its antigenicity score was 1.425 9 and immunogenicity score was 0.173 83. This epitope is very useful in the preparation of a universal vaccine against CHIKV infection. Conclusions: Epitopes reported in this study showed promiscuity, antigenicity as well as good binding affinity for the HLA classes. These epitopes will provide the baseline for development of efficacious vaccine for CHIKV.

Identification of an HLA-A~* 0201-restricted CD8^+ T-cell epitope SSp-1 of SARS-CoV spike protein

A novel coronavirus, severe acute respiratory syndrome (SA RS)-associated coronavirus (SARS-CoV), has been identified as the causal agent of SARS. Spike (S) protein is a major structural glycoprotein of the SARS virus and a potential target for SARS-specific cell-mediated immune responses. A pa nel of S protein-derived peptides was tested for their binding affinity to HLA -A *0201 molecules. Peptides with high affinity for HLA-A *0201 were then as se ssed for their capacity to elicit specific immune responses mediated by cytotoxi c T lymphocytes (CTLs) both in vivo, in HLA-A2.1/K b transgenic mice, a nd in vitro, from peripheral blood lymphocytes (PBLs) harvested from healthy HLA-A 2.1 + donors. SARS-CoV protein-derived peptide-1 (SSp-1 RLNEVAKNL), induced pepti de-specific CTLs both in vivo (transgenic mice) and in vitro (human PBL s), which specifically released interferon-gamma (IFN-gamma) upon stimulation with SSp-1-pulsed autologous dendritic cells (DCs) or T2 cells. SSp-1-specif ic CTLs also lysed major histocompatibility complex (MHC)-matched tumor cell lines engineered to express S proteins. HLA-A *0201-SSp-1 tetramer staining re vealed the presence of significant populations of SSp-1-specific CTLs in SSp- 1-induced CD8 + T cells. We propose that the newly identified epitope SSp-1 w ill help in the characterization of virus control mechanisms and immunopathology in SARS-CoV infection, and may be relevant to the development of immunotherape utic approaches for SARS.

Identification of the epitopes on HCV core protein recognized by HLA-A2 restricted cytotoxic T lymphocytes

AIM To identify hepatitis C virus (HCV) core protein epitopes recognized by HLA-A2 restricted cytotoxic T lymphocyte (CTL).METHODS Utilizing the method of computer prediction followed by a 4 h 51 Cr-release assay confirmation.RESULTS The results showed that peripheral blood mononuclear cells (PBMC) obtained from two HLA-A2 positive donors who were infected with HCV could lyse autologous target cells labeled with peptide 'ALAHGVFAL (core TS0-158)'.The rates of specific lysis of the cells from the two donors were 37.5% and 15.8%,respectively. Blocking of the CTL response with anti-CD4 mAb caused no significant decrease of the specific lysis.But blocking of CTL response with anti-CD8 mAb could abolish the Iysis.CONCLUSION The peptide (core 150 - 158 ) is the candidate epitope recognized by HLA-A2 restricted CTL.

Catalytic domain of PDC-E2 contains epitopes recognized by antimitochondrial antibodies in primary biliary cirrhosis

AIM:To search for further immunodominant peptides of the pyruvate dehydrogenase complex E2-component (PDC-E2) recognized by antimitochondrial antibodies (AMA) in primary biliary cirrhosis (PBC). METHODS:Sera from 95 patients with PBC were tested by enzyme-linked immunosorbent assay against 33 synthetic overlapping peptides (25 amino acids; aa) covering the entire length of the E2-subunit of PDC-E2. Furthermore,the inner lipoyl peptide 167-184 was used in an unlip oylated and a lipoylated form as well as coupled to ovalbumin. Sera from 11 AMA negative/ANA posit ive PBC patients,63 patients with other liver disorders and 22 healthy blood donors served as controls.RESULTS:Of the 95 PBC-sera,74% reacted with the peptide 475-499 and 58% with the pept ide 407-431 located within the catalytic domain of PDC-E2. Patients with other disorders or healthy controls were positive in only up to 18%. Antibodies to the unlipoylatedand lip oylated pept ide 167-184 within the inner lipoyl domain were found in only 5% and 11% of the PBC sera,respectively; using ovalbumin-coupled peptides,the incidence increased up to 57% (unlipoylated form). CONCLUSION:Peptides within the catalytic site of PDC-E2 rather than the previously reported lipoyl binding peptide 167-184 may represent major immunodomin ant epitopes recognized by AMA in PBC.

Bioinformatics analysis of the structure and linear B-cell epitopes of aquaporin-3 from Schistosoma japonicum

Objective:To analyze the structure of aquaporins-3(AQP-3) from Schistosoma japonicum(SJAQP-3) using bioinformalical methods,and to provid of references for vaccine targets research.Methods:Protparam,BepiPred,TMHMM Server,MLRC,Geno3d,DNA star software packages were used to predict the physical and chemical properties,hydrophilicity plot, flexibility regions,antigenic index,surface probability plot,secondary structure,and tertiary structure of amino acid sequence of SJAQP-3.Results:SJAQP-3 had six transmembrane regions and two half-spanning helices that form a central channel.The half-spanning helices fold into the centre of the channel.Either of the half-spanning helix had a conserved motif of NPA common to all aquaporins.Predicted linear B-Cell epitopes were most likely at the N-terminal amino acid residues of Saa-7aa,59aa- 62aa,225aa-230aa,282aa -288aa,294aa -29Saa and 305aa -307aa area.59aa- 62aa,22Saa-230aa located outside the membrane,the others located inside the cell.Conclusions:SJAQP-3 is a integral membrane protein in Schistosoma japonicum tegument.There are six potential epitopes in SJ AQP-3.It might be a potential molecular target for the development of vaccines.

Hepatocellular carcinoma-specific immunotherapy with synthesized α1,3-galactosyl epitope-pulsed dendritic cells and cytokine-induced killer cells

AIM: To evaluate the safety and clinical efficacy of a new immunotherapy using both α-Gal epitope-pulsed dendritic cells (DCs) and cytokine-induced killer cells.METHODS: Freshly collected hepatocellular carcinoma(HCC) tumor tissues were incubated with a mixture of neuraminidase and recombinant α1,3-galactosyltransferase (α1,3GT) to synthesize α-Gal epitopes on carbohydrate chains of the glycoproteins of tumor membranes. The subsequent incubation of the processed membranes in the presence of human natural anti-Gal IgG resulted in the effective phagocytosis to the tumor membrane by DCs. Eighteen patients aged 38-78 years with stage Ⅲ primary HCC were randomLy chosen for the study; 9 patients served as controls, and 9 patients were enrolled in the study group.RESULTS: The evaluation demonstrated that the procedure was safe; no serious side effects or autoimmune diseases were observed. The therapy significantly prolonged the survival of treated patients as compared with the controls (17.1 ± 2.01 mo vs 10.1 ± 4.5 mo,P = 0.00121). After treatment, all patients in the study group had positive delayed hyper sensitivity and robust systemic cytotoxicity in response to tumor lysate as measured by interferon-γ-expression in peripheral blood mononuclear cells using enzyme-linked immunosorbent spot assay. They also displayed increased numbers of CD8-, CD45RO-and CD56-positive cells in the peripheral blood and decreased α-fetoprotein level in the serum.CONCLUSION: This new tumor-specific immunotherapy is safe, effective and has a great potential for the treatment of tumors.

Preparation and Initial Application of a Monoclonal Antibody Specific for a Newly Discovered Conserved Linear Epitope of Rabies Virus Nucleoprotein

Objective To prepare monoclonal antibodies against a newly discovered and conserved linear epitope of Rabies virus nucleoprotein and to use them in a rabies diagnostic test. Methods Synthetic peptide containing the epitope was used as immunogen to prepare hybridoma cell lines by classical hybridoma technology. Anti-peptide monoclonal antibodies produced in ascites of inoculated Balb/c mice were labeled with fluorescein isothiocyanate （FITC） after purification and used in fluorescent antibody test （FAT）. Results Two positive hybridoma cell lines, RVNP-mAbl-CL and RVNP-mAb2-CL, were obtained. RVNP- mAbl-CL produced a higher concentration of monoclonal antibody RVNP-mAbl in Balb/c ascites. FITC-labeled RVNP-mAbl showed correct results on certain Rabies virus-positive canine brain tissue samples and cells of a small subclone of baby hamster kidney 21 cell line （BSR）. Conclusion FITC-labeled RVNP-mAbl has potential application for laboratory diagnosis of rabies

An immunoglobulin Y that specifically binds to an in silico-predicted unique epitope of Zika virus non-structural 1 antigen

Objective:To identify unique immunogenic epitopes of Zika virus non-structural 1(NS1)antigen and produce immunoglobulin Y(IgY)for potential use in he diagnosis of of Zika virus infection.Methods:Immunogenic epitopes were identified using in silico B-cell epitope prediction.A synthetic peptide analog of the predicted epitope was used to induce antipeptide IgY production in hens which was purified using affinity chromatography.Presence of purified IgY and its binding specificity were performed by gel electrophoresis and ELISA,respectively.Results:Out of the nine continuous epitopes identified,the sequence at position 193-208(LKVREDYSLECDPAVI)was selected and used to produce anti-peptide IgY.The produced IgY was found to bind to the synthetic analog of the Zika virus NS1 immunogenic epitope but not to other flaviviruses and random peptides from other pathogens.Conclusions:In this study,we identified an immunogenic epitope unique to Zika virus that can be used to develop a serodiagnostic tool that specifically detect Zika virus infection.

Epitope finding in Zika virus molecule:The first world report

Zika virus infection is a new problematic virus infection that becomes the present public health problem. Now this mosquito borne infectious disease can be seen worldwide and can cause dengue-like infection. In addition, it can also induce transplacental infection and result in congenital neurological defect. To prevent this infection, there is still no specific vaccine. To find a new vaccine, finding epitope is the first step. Here, the authors report the study to find epitope within Zika virus molecule.According to this study, the appropriate epitopes can be seen. This is the fi rst world report on epitope finding for Zika virus. The data can be useful for further vaccine development.

Prediction of T cell and B cell epitopes of the 22-, 47-, 56-, and 58-kDa proteins of Orientia tsutsugamushi

Objective:To predict B cell and T cell epitopes of 22-kDa,47-kDa,56-kDa and 58-kDa proteins.Methods:The sequences of 22-kDa,47-kDa,56-kDa and 58-kDa proteins which were derived from Orientia tsutsugamushi were analyzed by SOPMA,DNAstar,Bcepred,ABCpred,NetMHC,NetMHCⅡand IEDB.The 58-kDa tertiary structure model was built by MODELLER9.17.Results:The 22-kDa B-cell epitopes were located at positions 194-200,20-26 and 143-154,whereas the T-cell epitopes were located at positions 154-174,95-107,17-25 and 57-65.The 47-kD a protein B-cell epitopes were at positions 413-434,150-161 and 283-322,whereas the T-cell epitopes were located at positions 129-147,259-267,412-420 and 80-88.The 56-kDa protein B-cell epitopes were at positions 167-173,410-419 and 101-108,whereas the T-cell epitopes were located at positions 88-104,429-439,232-240 and 194-202.The 58-kDa protein B-cell epitopes were at positions 312-317,540-548 and 35-55,whereas the T-cell epitopes were located at positions 415-434,66-84 and 214-230.Conclusions:We identified candidate epitopes of 22-kDa,47-kDa,56-kDa and 58-kDa proteins from Orientia tsutsugamushi.In the case of 58-kDa,the dominant antigen is displayed on tertiary structure by homology modeling.Our findings will help target additional recombinant antigens with strong specificity,high sensitivity,and stable expression and will aid in their isolation and purification.

Natural epitope variants of the hepatitis C virus impair cytotoxic T lymphocyte activity

AIM:To understand how interactions between hepatitis C virus(HCV) and the host's immune system might lead to viral persistence or effective elimination of HCV.METHODS:Nucleotides 3519-3935 of the non-structural 3(NS3) region were amplified by using reverse transcription polymerase chain reaction(PCR).PCR products of the HCV NS3 regions were integrated into a PCR T7TOPO TA vector and then sequenced in both directions using an automated DNA sequencer.Relative major histocompatibility complex binding levels of wild-type and variant peptides were performed by fluorescence polarization-based peptide competition assays.Peptides with wild type and variant sequences of NS3 were synthesized locally using F-moc chemistry and purified by high-performance liquid chromatography.Specific cytotoxic T lymphocytes(CTLs) clones toward HCV NS3 wild-type peptides were generated through limiting dilution cloning.The CTL clones specifically recognizing HCV NS3 wild-type peptides were tested by tetramer staining and flow cytometry.Cytolytic activity of CTL clones was measured using target cells labeled with the fluorescence enhancing ligand,DELFIA EuTDA.RESULTS:The pattern of natural variants within three human leukocyte antigen(HLA)-A2-restricted NS3 epitopes has been examined in one patient with chronic HCV infection at 12,28 and 63 mo post-infection.Results obtained may provide convincing evidence of immune selection pressure for all epitopes investigated.Statistical analysis of the extensive sequence variation found within these NS3 epitopes favors a Darwinian selection model of variant viruses.Mutations within the epitopes coincided with the decline of CTL responses,and peptide-binding studies suggested a signif icant impact of the mutation on T cell recognition rather than peptide presentation by HLA molecules.While most variants were either not recognized or elicited low responses,such could antagonize CTL responses to target cells pulsed with wild-type peptides.CONCLUSION:Cross-recognition of CTL epitopes from wild-type and naturally-occurring HCV variants may lead to impaired immune responses and ultimately contribute to viral persistence.

Epitope Analysis of Anti-SARS-CoV-2 Neutralizing Antibodies

Coronavirus disease 2019 is threatening thousands of millions of people around the world.In the absence of specific and highly effective medicines,the treatment of infected persons is still very challenging.As therapeutics,neutralizing antibodies(NAbs)have great potential.Many NAbs have been reported,and most target various regions on the receptor-binding domain of the spike(S)protein,or the N-terminal domain.Several NAbs and NAb cocktails have been authorized for emergency use,and more arc in clinical trials or are under development.In this review,considering the angle of binding epitopes on the S protein,we summarize the functions and the underlying mechanisms of a set of well-recognized NAbs and provide guidance for vaccine design and the combinatorial use of these antibodies.In addition,we review the NAbs and NAb cocktails that have been approved for emergency use and discuss the effectiveness of these NAbs for combating severe acute respiratory syndrome coronavirus 2 mutants.

GPC3 fused to an alpha epitope of HBsAg acts as an immune target against hepatocellular carcinoma associated with hepatitis B virus

BACKGROUND:The incidence of hepatocellular carcinoma (HCC)in China is closely related to the population infected with hepatitis B virus(HBV).HCC cells with HBV secrete soluble HBsAg into blood but do not express it on the cell membrane This study aimed to construct and investigate a new glycosyl phosphatidylinositol(GPI)-anchored protein(GPC3+α+EGFP) as a DNA vaccine against HCC associated with HBV. METHODS:A recombinant plasmid(pcDNA3.1(+)/GPC3+ α+EGFP)was constructed and verified by restriction endo nuclease digestion and sequencing.pcDNA3.1(+)/GPC3+α+ EGFP was transfected into HepG2 cells(experimental group) using lipofectamine 2000.pEGFP-N1-transfected HepG2 cells were used as a negative control,and non-transfected HepG2 cells sreved as a blank control.HepG2 cells that steadily expressed the fusion protein GPC3+α+EGFP were screened by G418,propagated,and co-cultured with lymphocytes from healthy donors.Cell proliferation was measured by the classic sulforhodamine B assay.Apoptosis was assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL),and Fas gene transcription was determined by quantitative fluorescent PCR. RESULTS:The pcDNA3.1(+)/GPC3+α+EGFP plasmid was successfully constructed.In the experimental group,green fluorescence was observed at the cell periphery and in the cytoplasm,whereas in the negative control group,fluorescence was evenly distributed throughout the cell.Proliferation of the experimental group significantly decreased after 72 hours compared to the negative and blank control groups.Furthermore,the number of apoptotic cells was statistically different among the three groups as determined by a contingency table Chisquare test;the experimental group had the highest incidence of apoptosis.Fas gene transcription in the experimental group was higher than in the two control groups,and an increasing trend with time in the experimental group was observed. CONCLUSION:A chimeric,membrane-anchored protein, GPC3+α+EGFP,localized to the membrane of HepG2 cells and inhibited proliferation and accelerated apoptosis through a Fas-FasL pathway after co-cultivation with lymphocytes.

MAPPING EPITOPES OF HUMAN PAPILLOMAVIRUS TYPE 16 L1 PROTEIN WITH A PHAGE DISPLAY EPITOPE LIBRARY

The objective of this study is to map epitopes on HPMAPV16 L1 protein and provide information to the design of HPV16 prophylactic peptide vaccine. The epitopes on L1 protein were screenedby polyclonal and two monoclonal antibodies (BS and F4G3) against RPV16 L1 protin from a 6-merfd phage display epitope library with the method or immuuo-afrinity screening (Biopauuing). Aferthree rounds or Bio-Panning, the Positive phages were detected by L1 antibodies again with ELISA.The positive phages reacted strongly with L1 antibodies were then identified by DNA sequencing.Three mimotopes have been screened by polycloual and two monoclonal antibodies. The mimotope(LSLFSC) reacted with mouoclonal antibody B8 showed 50% pomology with the sequence 270275a. a (DSLFFY) of prototype HPV16 L1. Another mimotope (LTSSYS) reacted with polyclonalantibodies had 66% pomology with the L1 sequence 516~521a. A(TTSSTS), also a mimotope (DRWDRF) was found had the bomologic RF with the known L1 sequence 441 ~446a. a. The mimotopesLSLFSC and DRWDRF were adjacent to the epitopes at 267~269a. a and 422~441 a. a reported byother researchers Previously. Our results suggest that there might be a batch of epitopes on HPV16L1 ppotein, and the predominant epitopes of HPV16 L1 protein are located in the above two domains. These results will be helpful for design or HPV16 prophylactic peatide vaccines and HPVpolyvalent vaccines.