Functional analyses of Plasmodium Falciparum primary metabolic genes

Exploitation of plasmodial genes as drug-targets is important but their primary metabolic genes have been neglected and widely thought to be cryptic. However, datamining the plasmodial genome revealed DNA sequences that potentially support a network of glycolytic and TCA pathways. To question the existence of these genes, 10 predicted ORFs were cloned and expressed in Escherichia coli. This resulted in the soluble expression, authentication and kinetic characterization of P. falciparum isocitrate dehydrogenase, malate dehydrogenase and pyruvate kinase. And, since RT-PCR showed that all the 10 ORFs were actively transcribed during infection, analyses of web-based microarray data were done. These further supported the importance of searching transcription data to correlate genome annotations. To solve the need for a novel yet simple protocol to position plasmodial cellular genes, in-situ localization of fluorescently-labeled probes was developed and used to locate mitochondrial-, nuclear-, and apicoplast-targeting signal peptides in CHO-K1 cells. Taken together, this study has validated a plausible interactive network of primary metabolic plasmodial genes.

Author dc:creator

• CHAN KOK LEONG, MAURICE

• Glycolysis; tricarboxylic acid cycle; malaria genes; heterologous expression; transfection; localization