Department of Biological Sciences
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Item Plasmodium vivax apicoplast genome: A comparative analysis of major genes from Indian field isolates(Elsiever, 2012-04) Garg, Shilpi; Saxena, Vishal; Das, AshisThe apicomplexan parasite Plasmodium vivax is responsible for causing more than 70% of human malaria cases in Central and South America, Southeastern Asia and the Indian subcontinent. The rising severity of the disease and the increasing incidences of resistance shown by this parasite towards usual therapeutic regimens have necessitated investigation of putative novel drug targets to combat this disease. The apicoplast, an organelle of procaryotic origin, and its circular genome carrying genes of possible functional importance, are being looked upon as potential drug targets. The genes on this circular genome are believed to be highly conserved among all Plasmodium species. Till date, the plastid genome of P. falciparum, P. berghei and P. chabaudi have been detailed while partial sequences of some genes from other parasites including P. vivax have been studied for identifying evolutionary positions of these parasites. The functional aspects and significance of most of these genes are still hypothetical. In one of our previous reports, we have detailed the complete sequence, as well as structural and functional characteristics of the Elongation factor encoding tufA gene from the plastid genome of P. vivax. We present here the sequences of large and small subunit rRNA (lsu and ssu rRNA) genes, sufB (ORF470) gene, RNA polymerase (rpo B, C) subunit genes and clpC (casienolytic protease) gene from the plastid genome of P. vivax. A comparative analysis of these genes between P. vivax and P. falciparum reveals approximately 5–16% differences. A codon usage analysis of major plastid genes has shown a high frequency of codons rich in A/T at any or all of the three positions in all the species. TTA, AAT, AAA, TAT, and ATA are the major preferred codons. The sequences, functional domains and structural analysis of respective proteins do not show any variations in the active sites. A comparative analysis of these Indian P. vivax plastid genome encoded genes has also been done to understand the evolutionary position of the Indian parasite in comparison to other Plasmodium species.Item Structural and functional characterization of an iron–sulfur cluster assembly scaffold protein-SufA from Plasmodium vivax(Elsiever, 2016-07-01) Garg, Shilpi; Yadav, Sushil Kumar; Saxena, VishalIron–sulfur (Fe–S) clusters are utilized as prosthetic groups in all living organisms for diverse range of cellular processes including electron transport in respiration and photosynthesis, sensing of ambient conditions, regulation of gene expression and catalysis. In Plasmodium, two Fe–S cluster biogenesis pathways are reported, of which the Suf pathway in the apicoplast has been shown essential for the erythrocytic stages of the parasite. While the initial components of this pathway detailing the sulfur mobilization have been elucidated, the components required for the assembly and transfer of Fe–S clusters are not reported from the parasite. In Escherichia coli, SufB acts as a scaffold protein and SufA traffics the assembled Fe–S cluster from SufB to target apo-proteins. However, in Plasmodium, the homologs of these proteins are yet to be characterized for their function. Here, we report a putative SufA protein from Plasmodium vivax with signature motifs of A-type scaffold proteins, which is evolutionarily conserved. The presence of the [Fe4S4]3 + cluster under reduced conditions was confirmed by UV–visible and EPR spectroscopy and the interaction of these clusters with the conserved cysteine residues of chains A and B of PvSufA, validates its existence as a dimer, similar to that in E. coli. The H-bond interactions at the PvSufA–SufB interface demonstrate SufA as a scaffold protein in conjunction with SufB for the pre-assembly of Fe–S clusters and their transfer to the target proteins. Co-localization of the protein to the apicoplast further provides an experimental evidence of a functional scaffold protein SufA for the biogenesis of Fe–S clusters in apicoplast of PlasmodiumItem Characterization of 4-hydroxy-3-methylbut-2-en-1-yl diphosphate synthase (IspG) from Plasmodium vivax and it’s potential as an antimalarial drug target(Elsiever, 2017-03) Garg, Shilpi; Yadav, Sushil Kumar; Saxena, VishalThe prokaryotic type Methyl Erythritol phosphate (MEP) pathway functional in the apicoplast of Plasmodium is indispensable for the erythrocytic stages of the parasite. It is the sole process of isoprenoids biosynthesis in the parasite and is different from that in humans. Among the seven enzymes known to be functional in the MEP pathway in prokaryotes, most enzymes from Plasmodium are yet uncharacterized. The penultimate enzyme of this pathway 4-hydroxy-3-methylbut-2-en-1-yl diphosphate synthase (IspG), has been shown to act as a key target molecule in prokaryotes, where its deletion results in impairment of many housekeeping functions. The present study is the first detailed report of IspG enzyme from any Plasmodium sp. We report here that the protein is highly conserved across apicomplexans and prokaryotes and it localizes to the apicoplast as evident from the immune-localization studies performed on P. vivax infected blood smears made from clinical patients. The biochemical reconstitution and in silico docking of [4Fe–4S] clusters on the protein indicate their importance for the activity of enzyme. In-silico screening of different drug entities suggested the inhibitory role of alkyne diphosphate analogues and fosmidomycin against the IspG enzyme, suggesting the potential role of this enzyme as an antimalarial target.Item Deciphering the role of IspD (2‑C‑methyl‑D‑erythritol 4‑phosphate cytidyltransferase) enzyme as a potential therapeutic drug target against Plasmodium vivax(Elsiever, 2018-10-30) Garg, Shilpi; Saxena, VishalIspD(2‑C‑methyl‑d-erythritol 4-phosphate cytidyltransferase) PvIspDPlasmodium vivax IspD PfIspDPlasmodium falciparum IspD AtIspDArabidopsis thaliana IspD MEPMethyl erythritol phosphate (Non-mevalonate) HTSHigh Throughput Screening NEATnuclear encoded apicoplast targeted CTPcytidine 5′-triphosphate CDP-ME4-diphosphocytidyl-2C-methyl‑D-erythritol ACDacid citrate dextrose IHECInstitute Human Ethics Committee PCRPolymerase chain reaction MBPMaltose binding protein RIG plasmidPlasmid with arginine (R), Isoleucine (I) and Glycine (G) tRNA genes DOPEDiscrete Optimized Protein Energy pMALMaltose FITCFluorescein isothiocyanate DAPI4′,6-diamidino-2-phenylindole Qdotquantum dot RMSDroot-mean-square deviation