Characterization of a glycerophosphodiester phosphodiesterase in the human malaria parasite Plasmodium falciparum

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dc.contributor.authorDenloye, Titilola Ifeomaen
dc.contributor.committeechairKlemba, Michaelen
dc.contributor.committeememberHelm, Richard F.en
dc.contributor.committeememberHernick, Marcyen
dc.contributor.committeememberDean, Dennis R.en
dc.contributor.committeememberLarson, Timothy J.en
dc.contributor.departmentBiochemistryen
dc.date.accessioned2017-04-06T15:42:36Zen
dc.date.adate2012-06-08en
dc.date.available2017-04-06T15:42:36Zen
dc.date.issued2012-04-25en
dc.date.rdate2016-09-27en
dc.date.sdate2012-05-16en
dc.description.abstractActive lipid metabolism is a key process required for the intra-erythrocytic development of the malaria parasite, Plasmodium falciparum. Enzymes that hydrolyze host-derived lipids play key roles in parasite growth, virulence, differentiation, cell-signaling and hemozoin formation. Therefore, investigating enzymes involved in lipid degradation could uncover novel drug targets. We have identified in P. falciparum, a glycerophosphodiester phosphodiesterase (PfGDPD), involved in the downstream pathway of phosphatidylcholine degradation. PfGDPD hydrolyzes deacylated phospholipids, glycerophosphodiesters to glycerol-3-phosphate and choline. In this study, we have characterized PfGDPD using bioinformatics, biochemical and genetic approaches. Knockout experiments showed a requirement for PfGDPD for parasite survival. Sequence analysis revealed PfGDPD possesses the unique GDPD insertion domain sharing a cluster of conserved residues present in other GDPD homologues. We generated yellow fluorescent fusion proteins that revealed a complex distribution of PfGDPD within the parasite cytosol, parasitophorous vacuole and food vacuole. To gain insight into the role of PfGDPD, sub-cellular localization was modulated and resulted in a shift in protein distribution, which elicited no growth phenotype. Kinetic analyses suggest PfGDPD activity is Mg₂⁺ dependent and catalytically efficient at the neutral pH environment of the parasitophorous vacuole. Next, our aim was to determine the upstream pathway that provides deacylated glycerophosphodiesters as substrate for PfGDPD. We identified via bioinformatics, a P. falciparum lysophospholipase (PfLPL1) that directly generates the substrate. Knockout clones were generated and genotyped by Southern and PCR analysis. The effects of PfLPL1 knockouts on parasite fitness were studied, and the results showed that PfLPL1was not required for parasite survival and proliferation.en
dc.description.degreePh. D.en
dc.identifier.otheretd-05162012-185144en
dc.identifier.sourceurlhttp://scholar.lib.vt.edu/theses/available/etd-05162012-185144/en
dc.identifier.urihttp://hdl.handle.net/10919/77090en
dc.language.isoen_USen
dc.publisherVirginia Techen
dc.rightsIn Copyrighten
dc.rights.urihttp://rightsstatements.org/vocab/InC/1.0/en
dc.subjectcholineen
dc.subjectlipid metabolismen
dc.subjectfatty aciden
dc.subjectmalariaen
dc.subjectglycerophosphodiester phosphodiesteraseen
dc.subjectPlasmodium falciparumen
dc.titleCharacterization of a glycerophosphodiester phosphodiesterase in the human malaria parasite Plasmodium falciparumen
dc.typeDissertationen
dc.type.dcmitypeTexten
thesis.degree.disciplineBiochemistryen
thesis.degree.grantorVirginia Polytechnic Institute and State Universityen
thesis.degree.leveldoctoralen
thesis.degree.namePh. D.en

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