Adoro, Stanley;Nwuba, Roseangela;Anumudu, Chiaka;Omosun, Yusuf;Nwagwu, Mark - Analysis of the genetic and amino acid sequence diversity of antigenic determinants in the Plasmodium falciparum C-terminal merozoite surface protein-1 encoding a 19-kDa protei

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Title: Analysis of the genetic and amino acid sequence diversity of antigenic determinants in the Plasmodium falciparum C-terminal merozoite surface protein-1 encoding a 19-kDa protein	P2
Adoro, Stanley; Nwuba, Roseangela; Anumudu, Chiaka; Omosun, Yusuf; Nwagwu, Mark stanleyadoro@hotmail.com Cellular Parasitology Programme, Department of Zoology, University of Ibadan, Ibadan, Nigeria

Comparison of variations in gene and amino acid sequences is important to reveal phylogeny, understand gene function conservation and provides a means for understanding the influence of molecular sequence evolution and diversity on immunogenic epitopes of target antigens. In this study, using bioinformatics and immunological techniques, we have compared the sequences of 22 database genetic datasets of the MSP119 gene region obtained from Plasmodium falciparum isolates and analyzed the influence of variations in these genes on epitopes of human antibodies. The gene sequence of Plasmodium falciparum merozoite surface protein-1 (MSP1), encoding a 195-kDa-membrane protein of malaria merozoites, has been divided into 17 blocks with different degrees of variation. Compared to others, block 17 is conserved and encodes the 19-kDa C-terminal product (MSP119) of the proteolytic processing of MSP1. MSP119 is required by merozoites for erythrocyte invasion (a critical step in malaria pathogenesis) and it is currently being evaluated as a malaria vaccine. Previous studies have shown that MSP119 is comprised of two epidermal growth factor (EGF)-like domains and that anti-MSP119 antibodies protect against malaria. We performed multiple sequence alignments and obtained significant identity in the 22 sequences analyzed. Four broad gene clusters were observed including MSP119 genes from the same regions with the exception of the Kenya isolates. Nucleotide substitutions resulting in amino acid variations were observed at 10 positions: 8 within the interconnecting loop regions and 2 within beta-sheet structures of the two EGF domains. Binding of human serum antibodies to mutant MSP119 with amino acid substitutions incorporated at various positions by site-directed mutagenesis in Western Blot assays were investigated in order to elucidate polyclonal antibody epitopes in natural malaria endemic populations. Amino acid substitutions were visualized on a model crystal structure of MSP119. Our data showed that substitutions, which affected the binding of the human antibodies, were located only in the beta-sheets of the first EGF-domain with the exception of a loop Cys12 to Ile mutation, which forms a sulphur bridge. These results support previous findings that showed a strong antibody response to the first EGF-domain of MSP119; it is indeed conceivable that the conserved EGF-domains should elicit a greater antibody response than the more variable loops. Variability in loop residues however may indicate the redundancy of these regions in MSP119 activity or the operation of positive selection pressures in this gene region that are advantageous to the parasite. Thus, using models of codon substitutions, it will be useful to analyze selection pressures in MSP119 gene in future investigations. Our results underscore the importance of determining the MSP119 sequences from more geographical regions where malaria is endemic.

[1] Altschul, S. F., et al. 1997. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25:3389-3402.
[2] Chappel J.A and Holder A.A. 1993. Monoclonal antibodies that inhibit Plasmodium falciparum invasion in vitro recognize the first growth factor-like domain of merozoite surface protein-1. Mol Biochem Parasitol 60: 303-312.
[3] Hall, T.A. 1999. BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucl. Acids. Symp. Ser. 41:95-98.
[4] Holder A.A., et al. 1999. Merozoite surface protein 1, immune evasion, and vaccines against asexual blood stage malaria. Parassitologia 41:409-414.
[5] Morgan W.D. et al. 1999. Solution structure of an EGF module pair from the Plasmodium falciparum merozoite surface protein 1. J Mol Biol 209: 113-122.
[6] Nwuba R.I., et al. 2002. The human immune response to Plasmodium falciparum includes antibodies that inhibit merozoite surface protein-1 processing and blocking antibodies. Infect. Immun., in press.
[7] Rich S.M. and Ayala F.J. 2000. Population structure and recent evolution of Plasmodium falciparum. Proc Natl Acad Sci USA 97:6994-7001.
[8] Tanabe K., et al. 1987. Allelic dimorphism in a surface antigen of the malaria parasite Plasmodium falciparum. J Mol Biol 195: 273-287.
[9] Thompson J.D., et al. 1994. CLUSTALW: improving the sensitivity of progressive multiple sequence alignments through weighting, positions-specific gap penalties and weight matrix choice. Nucl Acids Res 22: 4673-4680.
[10] Uthaipibull C., et al. 2001. Inhibitory and Blocking monoclonal antibody epitopes on merozoite surface protein 1 of the malaria parasite Plasmodium falciparum. J. Mol. Biol. 307:1381-1394.