In silico analysis of chimeric TF, Omp31 and BP26 fragments of Brucella melitensis for development of a multi subunit vaccine candidate

Document Type : Original Article

Authors

1 Molecular Biology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran

2 Applied Microbiology Research Center, Baqiyatallah Medical Science University, Tehran, Iran

Abstract

Objective(s):Brucellosis, especially caused by Brucella melitensis, remains one of the most common zoonotic diseases worldwide with more than 500,000 human cases reported annually. The commonly used live attenuated vaccine in ovine brucellosis prophylaxis is B. melitensis Rev1. But due to different problems caused by the administration of this vaccine, a protective subunit vaccine against B. melitensis is strongly demanded. Brucella BP26, Omp31 and TF proteins have shown a considerable potential as protective antigens for brucellosis. Chimeric proteins carrying epitopes or adjuvant sequences increase the possibility of eliciting a broad cellular or humoral immune response. In silico tools are highly suited to study, design and evaluate vaccine strategies.
Materials and Methods: In this study, a synthetic chimeric gene, encoding TF, BP26 93-111 and Omp3148-74 was designed.In order to predict the 3D structure of protein, modeling was carried out.
Results:Validation results showed that 91.1% of residues lie in favored or additional allowed region of Ramachandran plot. The epitopes in the chimeric protein are likely to induce both the B-cell and T-cell mediated immune responses.
Conclusion: The chimeric protein may be used as multi subunit for development of Brucella vaccine candidates.

Keywords


1. Siadat SD, Sharifat Salmani A, Aghasadeghi MR. Brucellosis Vaccines: An Overview, Zoonosis: Dr Jacob Lorenzo-Morales 2012.
2. Cassataro J, Pasquevich K, Bruno L, Wallach JC, Fossati CA, Baldi PC. Antibody reactivity to Omp31 from Brucella melitensis in human and animal Amani et al In silico analysis of chimeric TF, Omp31 and BP26 infections by smooth and rough Brucellae. Clin Diagn Lab Immunol 2004; 11:111-114.
3. Yang Y, Yin J, Guo D, Lang X, Wang X. Immunization of mice with recombinant S-adenosyl-L-homocysteine hydrolase protein confers protection against Brucella melitensis infection. FEMS Immunol Med Microbiol 2011; 61:159-167.
4. Zhao Z, Li M, Luo D, Xing L, Wu S, Duan Y,et al. Protection of mice from Brucella infection by immunization with attenuated Salmonella enterica serovar typhimurium expressing A L7/L12 and BLS fusion antigen of Brucella. Vaccine 2009; 27:5214-5219.
5. Sciutto E, Toledo A, Cruz C, Rosas G, Meneses G, Laplagne D,et al. Brucella spp. lumazine synthase: a novel antigen delivery system. Vaccine 2005; 23:2784-2790.
6. Yang X, Hudson M, Walters N, Bargatze RF, Pascual DW. Selection of protective epitopes for Brucella melitensis by DNA vaccination. Infect Immun 2005; 73:7297-7303.
7. Cassataro J, Estein SM, Pasquevich KA, Velikovsky CA, de la Barrera S, Bowden R, et al. Vaccination with the recombinant Brucella outer membrane protein 31 or a derived 27-amino-acid synthetic peptide elicits a CD4+ T helper 1 response that protects against Brucella melitensis infection. Infect Immun 2005; 73:8079-8088.
8. Cloeckaert A, Debbarh HS, Zygmunt MS, Dubray G. Production and characterisation of monoclonal antibodies to Brucella melitensis cytosoluble proteins that are able to differentiate antibody responses of infected sheep from Rev. 1 vaccinated sheep. J Med Microbiol 1996; 45:206-213.
9. Qiu J, Wang W, Wu J, Zhang H, Wang Y, Qiao J, et al. Characterization of periplasmic protein BP26 epitopes of Brucella melitensis reacting with murine monoclonal and sheep antibodies. PloS One 2012; 7:e34246.
10. Ghasemi A , Salari MH , Pourmand MR , Zarnani AH , Ahmadi H , Shirazi MH ,et al. Immune Reactivity of Brucella melitensis –Vaccinated rabbit serum with Recombinant Omp31 and DnaK proteins. Iran J Microbiol 2013; 5:19-23.
11. Cloeckaert A, Vizcaino N, Paquet JY, Bowden RA, Elzer PH. Major outer membrane proteins of Brucella spp.: past, present and future. Vet Microbiol 2002; 90:229-247.
12. Cassataro J, Pasquevich KA, Estein SM, Laplagne DA, Velikovsky CA, de la Barrera S, et al. A recombinant subunit vaccine based on the insertion of 27 amino acids from Omp31 to the N-terminus of BLS induced a similar degree of protection against B. ovis than Rev.1 vaccination. Vaccine 2007; 25:4437-4446.
13. Commander NJ, Spencer SA, Wren BW, MacMillan AP. The identification of two protective DNA vaccines from a panel of five plasmid constructs encoding Brucella melitensis 16M genes. Vaccine 2007; 25:43-54.
14. Luo D, Ni B, Li P, Shi W, Zhang S, Han Y, et al. Protective immunity elicited by a divalent DNA vaccine encoding both the L7/L12 and Omp16 genes of Brucella abortus in BALB/c mice. Infect Immun 2006; 74:2734-2741.
15. Araya LN, Winter AJ. Comparative protection of mice against virulent and attenuated strains of Brucella abortus by passive transfer of immune T cells or serum. Infect Immun 1990; 58:254-256.
16. Velikovsky CA, Goldbaum FA, Cassataro J, Estein S, Bowden RA, Bruno L, et al. Brucella lumazine synthase elicits a mixed Th1-Th2 immune response and reduces infection in mice challenged with Brucella abortus 544 independently of the adjuvant formulation used. Infect Immun 2003; 71:5750-5755.
17. Al-Mariri A, Tibor A, Mertens P, De Bolle X, Michel P, Godefroid J, et al. Protection of BALB/c mice against Brucella abortus 544 challenge by vaccination with bacterioferritin or P39 recombinant proteins with CpG oligodeoxynucleotides as adjuvant. Infect Immun 2001; 69:4816-4822.
18. Cespedes S, Andrews E, Folch H, Onate A. Identification and partial characterisation of a new protective antigen of Brucella abortus. J Med Microbiol 2000; 49:165-170.
19. Tabatabai LB, Pugh GW Jr. Modulation of immune responses in Balb/c mice vaccinated with Brucella abortus Cu-Zn superoxide dismutase synthetic peptide vaccine. Vaccine 1994; 12:919-924.
20. Oliveira SC, Splitter GA. Immunization of mice with recombinant L7/L12 ribosomal protein confers protection against Brucella abortus infection. Vaccine 1996; 14:959-962.
21. Maione D, Margarit I, Rinaudo CD, Masignani V, Mora M, Scarselli M, et al. Identification of a universal Group B streptococcus vaccine by multiple genome screen. Science 2005; 309:148-150.
22. Nazarian S, Mousavi Gargari SL, Rasooli I, Amani J, Bagheri S, Alerasool M. An in silico chimeric multi subunit vaccine targeting virulence factors of enterotoxigenic Escherichia coli (ETEC) with its bacterial inbuilt adjuvant. J Microbiol Methods 2012; 90:36-45.
23. Puigbo P, Guzman E, Romeu A, Garcia-Vallve S. OPTIMIZER: a web server for optimizing the codon usage of DNA sequences. Nucleic Acids Res 2007; 35:W126-31.
24. Amani J, Mousavi SL, Rafati S, Salmanian AH. In silico analysis of chimeric espA, eae and tir fragments of Escherichia coli O157:H7 for oral immunogenic applications. Theor Biol Med Model 2009; 6:28.
25. Keramati M, Roohvand F, Aslani MM, Khatami S, Aghasadeghi MR, Sadat M,et al. Screening, cloning and expression of active streptokinase from an Iranian isolate of S. equisimilis Group C in E. coli Iran J Basic Med Sci 2013; 16:620-627.
26. Doytchinova IA, Flower DR. VaxiJen: a server for prediction of protective antigens, tumour antigens and subunit vaccines. BMC Bioinformatics 2007; 8:4.
27. Iman M, Saadabadi A, Davood A. Docking studies of phthalimide pharmacophore as a sodium channel blocker. Iran J Basic Med Sci 2013; 16:1016-1021.
28. Rahbar MR, Rasooli I, Gargari SL, Sandstrom G, Amani J, Fattahian Y,et al. A potential in silico antibody-antigen based diagnostic test for precise identification of Acinetobacter baumannii. J Theor Biol 2011; 294:29-39. In silico analysis of chimeric TF, Omp31 and BP26 Amani et al
29. Ansari HR, Raghava GP. Identification of conformational B-cell Epitopes in an antigen from its primary sequence. Immunome Res 2010; 6:6.
30. Negi SS, Braun W. Automated detection of conformational epitopes using phage display Peptide sequences. Bioinformatics Biol Insights 2009; 3:71-81.
31. Singh H, Raghava GP. ProPred1: prediction of promiscuous MHC Class-I binding sites. Bioinformatics 2003; 19:1009-1014.
32. Singh H, Raghava GP. ProPred: prediction of HLA-DR binding sites. Bioinformatics 2001; 17:1236-1237.
33. Guan P, Doytchinova IA, Zygouri C, Flower DR. MHCPred: A server for quantitative prediction of peptide-MHC binding. Nucleic Acids Res 2003; 31:3621-3624.
34. Hamada M, Kiryu H, Sato K, Mituyama T, Asai K. Prediction of RNA secondary structure using generalized centroid estimators. Bioinformatics 2009; 25:465-473.
35. Sen TZ, Jernigan RL, Garnier J, Kloczkowski A. GOR V server for protein secondary structure prediction. Bioinformatics 2005; 21:2787-2788.
36. Rost B, Liu J. The PredictProtein server. Nucleic Acids Res 2003; 31:3300-3304.
37. Edwards YJ, Cottage A. Bioinformatics methods to predict protein structure and function. A practical approach. Mol Biotechnol 2003; 23:139-166.
38. Wiederstein M, Sippl MJ. ProSA-web: interactive web service for the recognition of errors in three-dimensional structures of proteins. Nucleic Acids Res 2007; 35:W407-410.
39. Laskowski RA, Rullmannn JA, MacArthur MW, Kaptein R, Thornton JM. AQUA and PROCHECK-NMR: programs for checking the quality of protein structures solved by NMR. J Biomol NMR 1996; 8:477-486.
40. Christen M, Hunenberger PH, Bakowies D, Baron R, Burgi R, Geerke DP, et al. The GROMOS software for biomolecular simulation: GROMOS05. J Comput Chem 2005; 26:1719-1751.
41. Ivanciuc O, Schein CH, Braun W. SDAP: database and computational tools for allergenic proteins. Nucleic Acids Res 2003; 31:359-362.
42. Zuker M. Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res 2003; 31:3406-3415.
43. Zhang Y. I-TASSER server for protein 3D structure prediction. BMC Bioinformatics 2008; 9:40.
44. Rost B, Sander C. Conservation and prediction of solvent accessibility in protein families. Proteins 1994; 20:216-226.
45. Cassataro J, Velikovsky CA, de la Barrera S, Estein SM, Bruno L, Bowden R, et al. A DNA vaccine coding for the Brucella outer membrane protein 31 confers protection against B. melitensis and B. ovis infection by eliciting a specific cytotoxic response. Infect Immun 2005; 73:6537-6546.
46. Delpino MV, Estein SM, Fossati CA, Baldi PC, Cassataro J. Vaccination with Brucella recombinant DnaK and SurA proteins induces protection against Brucella abortus infection in BALB/c mice. Vaccine 2007; 25:6721-6729.
47. Al-Mariri A. Protection of BALB/c mice against Brucella melitensis 16 M infection induced by vaccination with live Escherchia coli expression Brucella P39 protein. Vaccine 2010; 28:1766-1770.
48. Ferbitz L, Maier T, Patzelt H, Bukau B, Deuerling E, Ban N. Trigger factor in complex with the ribosome forms a molecular cradle for nascent proteins. Nature 2004; 431:590-596.
49. Deuerling E, Patzelt H, Vorderwulbecke S, Rauch T, Kramer G, Schaffitzel E,et al. Trigger factor and DnaK possess overlapping substrate pools and binding specificities. Mol Microbiol 2003; 47:1317-1328.
50. Debbarh HS, Zygmunt MS, Dubray G, Cloeckaert A. Competitive enzyme-linked immunosorbent assay using monoclonal antibodies to the Brucella melitensis BP26 protein to evaluate antibody responses in infected and B. melitensis Rev.1 vaccinated sheep. Vet Microbiol 1996; 53:325-337.
51. Seco-Mediavilla P, Verger JM, Grayon M, Cloeckaert A, Marin CM, Zygmunt MS, et al. Epitope mapping of the Brucella melitensis BP26 immunogenic protein: usefulness for diagnosis of sheep brucellosis. Clin Diagn Lab Immunol 2003; 10:647-651.
52. Cloeckaert A, Baucheron S, Vizcaino N, Zygmunt MS. Use of recombinant BP26 protein in serological diagnosis of Brucella melitensis infection in sheep. Clin Diagn Lab Immunol 2001; 8:772-775.
53. Estein SM, Cassataro J, Vizcaino N, Zygmunt MS, Cloeckaert A, Bowden RA. The recombinant Omp31 from Brucella melitensis alone or associated with rough lipopolysaccharide induces protection against
54. Arai R, Ueda H, Kitayama A, Kamiya N, Nagamune T. Design of the linkers which effectively separate domains of a bifunctional fusion protein. Protein Eng 2001; 14:529-532.