Hemozoin-free Plasmodium falciparum mitochondria for physiological and drug susceptibility studies.
Isolation of mitochondria of high purity and with intact enzymatic activities from malaria parasites has proven to be a major obstacle in characterizing the parasite mitochondrial physiology. We describe here an improved procedure for the isolation of a mitochondrially enriched preparation from the trophozoite stage of erythrocytic Plasmodium falciparum, combining disruption by N(2) cavitation and differential centrifugation with magnetic removal of hemozoin-associated material. These mitochondrial preparations may be used to assay various mitochondrial enzyme activities, such as succinate and dihydroorotate dehydrogenases, ubiquinol-cytochrome c oxidoreductase, and cytochrome c oxidase. They also exhibit a low level of ATPase activity, which is only marginally inhibited by classical inhibitors. We have used this preparation to determine the susceptibility of mitochondrial activities to drugs and drug candidate compounds in both "wild type" and transgenic parasites.
Mather
Michael W
MW
Center for Molecular Parasitology, Department of Microbiology and Immunology, Drexel University College of Medicine, Philadelphia, PA 19129, USA. mmather@drexelmed.edu
Morrisey
Joanne M
JM
Vaidya
Akhil B
AB
D000818:Animals; D000962:Antimalarials / Q000494:pharmacology*; D002458:Cell Fractionation; D000090722:Dihydroorotate Dehydrogenase; D014450:Electron Transport Complex III / Q000187:drug effects / Q000378:metabolism*; D004912:Erythrocytes / Q000469:parasitology; D006420:Hemeproteins / Q000737:chemistry; D006801:Humans; D008280:Magnetics; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000187:drug effects / Q000378:metabolism*; D021261:Parasitic Sensitivity Tests; D010274:Parasitology / Q000379:methods; D010963:Plasmodium falciparum / Q000187:drug effects* / Q000201:enzymology / Q000502:physiology* / Q000648:ultrastructure; D013053:Spectrophotometry
D000962:Antimalarials; D000090722:Dihydroorotate Dehydrogenase; D006420:Hemeproteins; C020753:hemozoin; D044925:Oxidoreductases Acting on CH-CH Group Donors; D014450:Electron Transport Complex III