Chew, C.H. and Lim, Yvonne Ai Lian and Chua, Kek Heng (2017) Heterologous expression of Plasmodium vivax apical membrane antigen 1 (PvAMA1) for binding peptide selection. PeerJ, 5. e3794. ISSN 2167-8359, DOI https://doi.org/10.7717/peerj.3794.
Full text not available from this repository.Abstract
Background. Plasmodium is an obligate intracellular parasite. Apical membrane antigen 1 (AMA1) is the most prominent and well characterized malarial surface antigen that is essential for parasite-host cell invasion, i.e., for sporozoite to invade and replicate within hepatocytes in the liver stage and merozoite to penetrate and replicate within erythrocytes in the blood stage. AMA1 has long served as a potent antimalarial drug target and is a pivotal vaccine candidate. A good understanding of the structure and molecular function of this Plasmodium protein, particularly its involvement in host-cell adhesion and invasion, is of great interest and hence it offers an attractive target for the development of novel therapeutics. The present study aims to heterologous express recombinant Plasmodium AMA1 ectodomain of P. vivax (rPvAMA1) for the selection of binding peptides. Methods. The rPvAMA1 protein was heterologous expressed using a tag-free Profinity eXactTM system and codon optimized BL21-Codon Plus (DE3)-RIL Escherichia coli strain and further refolded by dialysis for renaturation. Binding peptides toward refolded rPvAMA1 were panned using a Ph.D.-12 random phage display library. Results. The rPvAMA1 was successfully expressed and refolded with three phagedisplayed dodecapeptides designated as PdV1 (DLTFTVNPLSKA), PdV2 (WHWSWWNPNQLT), and PdV3 (TSVSYINNRHNL) with affinity towards rPvAMA1 identified. All of them exhibited positive binding signal to rPvAMA1 in both direct phage assays, i.e., phage ELISA binding assay and Western blot binding assay. Discussion. Phage display technology enables the mapping of protein-protein interactions based on a simple principle that a library of phage particles displaying peptides is used and the phage clones that bind to the target protein are selected and identified. The binding sites of each selected peptides toward PvAMA1 (Protein Data Bank, PDB ID: 1W8K) were in silico predicted using CABS-dock web server. In this case, the binding peptides provide a valuable starting point for the development of peptidomimetic as antimalarial antagonists directed at PvAMA1.
| Item Type: | Article |
|---|---|
| Funders: | Ministry of Higher Education Malaysia: Fundamental Research Grant Scheme (FRGS) (Project no. FRGS/1/2015/SKK12/UNISZA/02/1), University of Malaya, Malaysia: Postgraduate Research Grant (Project no. PS155-2008C and PS251-2010A) |
| Uncontrolled Keywords: | Apical membrane antigen 1 (AMA1); Binding peptide; In silico peptide docking; Phage display; Plasmodium vivax; Recombinant protein expression |
| Subjects: | R Medicine |
| Divisions: | Faculty of Medicine |
| Depositing User: | Ms. Juhaida Abd Rahim |
| Date Deposited: | 12 Sep 2018 04:12 |
| Last Modified: | 24 Oct 2019 08:01 |
| URI: | http://eprints.um.edu.my/id/eprint/19208 |
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