In silico approach to ascertain the calcium dependent role of Plasmodium falciparum SERA5.

The P. falciparum serine repeat antigen (PfSERA5) is the most abundantly expressed protein in the parasitophorous vacuole during the asexual blood stage and serves as both drug and vaccine target. The processed central fragment (56 KDa) of PfSERA5 is implicated to play an important role in parasite exit (egress) during schizont rupture from erythrocytes. Structural characterization of its enzymatic domain supports protease-like function for this central domain. The understanding of exact functional role of PfSERA5 in parasite egress remains unconfirmed as recent studies also indicate an indispensable non-catalytic role for PfSERA5 putative enzyme domain in the blood stage. No structural insight into PfSERA5 prodomain is available. Structure prediction of PfSERA5 prodomain using in silico approach in our study, showed it to have structural similarity with calcium-binding proteins. An earlier observation of steep rise in intracellular calcium concentration as an important factor in egress makes the prodomain calcium-binding role significant. The implication of calcium on structure and activity of PfSERA5 putative enzyme domain is also unknown, and such information would aid to substantiating any calcium-dependent effects on PfSERA5. To understand this, we performed molecular dynamic (MD) simulation both in the presence and absence of calcium. MD results show secondary structure conformational differences in local regions of protein structure. Our results support calcium to be an important parameter for stability and function of PfSERA5. This computational assessment suggest a need to design future experiments like calcium-dependent inhibition studies to reveal exact functional role of PfSERA5 in parasite egress.

Plasmodium vivax: N-terminal diversity in the blood stage SERA genes from Indian isolates.

Worldwide malaria risk due to Plasmodium vivax makes development of vaccine against P. vivax, a high priority. Serine Repeat Antigen of P. vivax (PvSERA) is a multigene family of blood stage proteins with 12 homologues. Sequence diversity studies are important for understanding them as potential vaccine candidates. No information on N-terminal diversity of these genes is available in literature. In this paper, we evaluate the genetic polymorphism of N-terminal regions of the highly expressed member PvSERA4 and PvSERA5 genes from Indian field isolates. Our results show that PvSERA4 has deletions and insertions in Glutamine rich tetrameric repeat units contributing to its diversity. PvSERA5 also exhibits high genetic diversity with non-synonymous substitutions leading to identification of novel haplotypes from India. Our first report helps in elucidating the allelic variants of PvSERA genes in this region and contributes to evaluating their efficacy as vaccine candidates.

Genetic and structural characterization of PvSERA4: potential implication as therapeutic target for Plasmodium vivax malaria.

Plasmodium vivax malaria is geographically the most widely distributed and prevalent form of human malaria. The development of drug resistance by the parasite to existing drugs necessitates higher focus to explore and identify new drug targets. Plasmodial proteases have key roles in parasite biology and are involved in nutritional uptake, egress from infected reticulocytes, and invasion of the new target erythrocytes. Serine repeat antigens (SERA) of Plasmodium are parasite proteases that remain attractive drug targets and are important vaccine candidates due to their high expression profiles in the blood stages. SERA proteins have a unique putative papain-like cysteine protease motif that has either serine or cysteine in its active site. In P. vivax, PvSERA4 is the highest transcribed member of this multigene family. In this study, we have investigated the genetic polymorphism of PvSERA4 central protease domain and deduced its 3D model by homology modeling and also performed MD simulations to acquire refined protein structure. Sequence analysis of protease domain of PvSERA4 from Indian field isolates reveals that the central domain is highly conserved. The high sequence conservation of the PvSERA4 enzyme domain coupled with its high expression raises the possibility of it having a critical role in parasite biology and hence, being a reliable target for new selective inhibitor-based antimalarial chemotherapeutics. The 3D model showed the presence of an unusual antiparallel Beta hairpin motif between catalytic residues similar to hemoglobin binding motif of Plasmodial hemoglobinases. Our PvSERA4 model will aid in designing structure-based inhibitors against this enzyme.

Plasmodium vivax: C-terminal diversity in the blood stage SERA genes from Indian field isolates.

The burden of Plasmodium vivax malaria is huge in India, affecting a large population annually. Recent reports of P. vivax contributing to severe illness and death, makes vaccine research on P. vivax malaria, a high priority. Extent of sequence variation in antigen coding genes is known to be a major hurdle in vaccine initiatives against malaria. Serine repeat antigens of Plasmodium are promising asexual blood stage vaccine candidates against malaria and have been implicated to have a key role in merozoite invasion and egress. Among the P. vivax SERA proteins, SERA4 and SERA5 are the major transcribed members in erythrocytic stages, making them encouraging candidates to be explored for their polymorphism and vaccine potential. Earlier reports suggest that diversity in these PvSERA antigens is localized to the C-terminal region of the proteins. Hence, genetic diversity study of this region seems prudent. Moreover, as there are no reports available from India, the present study aims to investigate the polymorphism in the C-terminal region of two highly transcribed members PvSERA4 and PvSERA5 in Indian field isolates. Our result of PvSERA5 demonstrates extensive genetic diversity, with major deletions, insertions and SNPs and signifies the gene to be under positive selection. On the other hand, high sequence conservation was seen in the PvSERA4 C-terminal region in Indian field isolates which was contrasting to earlier report from Thailand where they have shown diversity. Research data showcased in this study will greatly aid in gaining better understanding of antigenic variations, immune mediated selection mechanisms and the functional significance of these two vivax proteins. This study also makes a striking contribution towards understanding the antigenic repertoire of PvSERA genes in Indian isolates.