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Title: Identification and Characterization of Promiscuous Peptides from Plasmodium falciparum antigens
Researcher: Singh,Mr Shakti Pal
Guide(s): Farooq, Dr Umar
Keywords: HLA
Plasmodium falciparum
promiscuous peptide
vaccine candidate antigen
University: Shoolini University of Biotechnology and Management Sciences
Completed Date: 08-01-2016
Abstract: Abstract newlinePlasmodium falciparum is the most virulent malaria parasite causing severe form of disease which is potentially fatal. Even though extensive progress has been achieved in understanding the immune mechanisms involved in the malarial infection, there is no effective vaccine that has been introduced in to clinical practice. The most effective vaccine against malaria should be one which can induce immune responses against enormous epitopes in context of predominantly occurring HLA alleles. Distinct binding chemistry of different peptides and HLA polymorphism are the major challenges in the development of new vaccine candidates. The use of bioinformatics tools, reduce the time and efforts involve in the screening of potential peptides which can bind to several human leukocyte antigens. The present study was aimed to identify the promiscuous peptide of potential vaccine candidate antigens like merozoite surface antigen 1 and 2 (MSP-1 and MSP-2), circumsporozoite protein (CSP), GLURP, EBA-175 and Soluble antigen (S-antigen) of P. falciparum by using sequence and structure based in silico method. The consensus approach was applied to the currently best available servers for prediction of promiscuous peptides. The predicted promiscuous peptides were further verified by structure based in-silico method (Molecular docking). Promiscuous peptides with minimum free binding energy were selected for further in-vitro analysis. Cells were cultured with different peptide concentrations (25and#61549;g/ml, 50and#61549;g/ml and 100and#61549;g/ml), phytohaemagglutinin (PHA) was used as positive control and without peptide as negative control. The cell acquisition was done by flow cytometer and analysis of the results was performed by FACSDIVA and flowjo V_10 software. Culture supernatants from stimulated human PBMC were collected and assayed for IL-4 and IFN-and#947; production using a specific enzyme-linked immunosorbent Assay (ELISA) kit. HLA typing was done by PCR-SSP low resolution HLA typing kit Innotrain Germany. newlineWe have predicted 1706 peptides for MSP-1, 258 peptides for MSP-2, 383 for CSP, 1488 for EBA-175, 571 for S-antigen and 1488 for GLURP antigen. On the basis of 50% and 100% binding affinity to HLA allele selection criteria, 14 promiscuous peptides for MSP-2, 05 for CSP and 09 for EBA-175 antigen were obtained. Further by using the criteria of strong binding affinity (IC50lt1000) 04 promiscuous peptides for MSP-2, 01 for CSP and 02 for EBA-175 were found strong binders promiscuous peptides. These peptides were further verified by docking analysis and the promiscuous peptide EBA-9 from RII region of EBA-175 was selected for synthesis on the basis of best dock confirmation and minimum binding energy (-1.95K cal/mol). newlineviii newlineThe peptide showed an encouraging immunological response and T-cell proliferation was observed highest with 100and#956;g/ml concentration while 50and#956;g/ml showed moderate proliferation and percentage immune response was 100% for exposed population. The PBMCs obtained from unexposed population showed very low proliferation of lymphocytes. The expression of IFN- and#61543;and#61472;was observed significantly high in supernatant of sensitized cultured PBMCs whereas the expression of IL-4 was observed very low. newlineThe present study also suggested that the in-silico technique is a powerful tool to develop a vaccine based on promiscuous peptides. While a positive immunogenic response against synthetic peptide predicted from RII region of EBA-175 antigen reveals that EBA-175 can generate T-cell lymphoproliferation and can be a part of cocktail vaccine against malaria. newlineKey words: vaccine candidate antigen, HLA, promiscuous peptide, in-silico, Plasmodium falciparum. newline
Pagination: viii,132
Appears in Departments:Faculty Of Biotechnology

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01cover page.pdfAttached File260.9 kBAdobe PDFView/Open
02-certificates.pdf282.88 kBAdobe PDFView/Open
03-contents.pdf108.77 kBAdobe PDFView/Open
04-acknowledgement.pdf153.72 kBAdobe PDFView/Open
05-abbrevation.pdf96.21 kBAdobe PDFView/Open
06-list of figures.pdf166.48 kBAdobe PDFView/Open
07-list of tables.pdf101.06 kBAdobe PDFView/Open
08-abstract.pdf171.48 kBAdobe PDFView/Open
09-introduction.pdf224.89 kBAdobe PDFView/Open
10-review of literature.pdf376.42 kBAdobe PDFView/Open
11-material and methods.pdf460.67 kBAdobe PDFView/Open
12-results.pdf1.41 MBAdobe PDFView/Open
13-discussion.pdf233.38 kBAdobe PDFView/Open
14-summary and conclusion.pdf128.09 kBAdobe PDFView/Open
15-refrences.pdf350.96 kBAdobe PDFView/Open
16-appendix.pdf2.36 MBAdobe PDFView/Open
publication-1.pdf76.46 kBAdobe PDFView/Open
publication-2.pdf76.52 kBAdobe PDFView/Open
publication-3.pdf111.04 kBAdobe PDFView/Open

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