title: Characterization of two mitochondrial flavin adenine dinucleotide-dependent glycerol-3-phosphate dehydrogenases in Trypanosoma brucei.
abstract: Glycerol-3-phosphate dehydrogenases (G3PDHs) constitute a shuttle that serves for regeneration of NAD(+) reduced during glycolysis. This NAD-dependent enzyme is employed in glycolysis and produces glycerol-3-phosphate from dihydroxyacetone phosphate, while its flavin adenine dinucleotide (FAD)-dependent homologue catalyzes a reverse reaction coupled to the respiratory chain. Trypanosoma brucei possesses two FAD-dependent G3PDHs. While one of them (mitochondrial G3PDH [mtG3PDH]) has been attributed to the mitochondrion and seems to be directly involved in G3PDH shuttle reactions, the function of the other enzyme (putative G3PDH [putG3PDH]) remains unknown. In this work, we used RNA interference and protein overexpression and tagging to shed light on the relative contributions of both FAD-G3PDHs to overall cellular metabolism. Our results indicate that mtG3PDH is essential for the bloodstream stage of T. brucei, while in the procyclic stage the enzyme is dispensable. Expressed putG3PDH-V5 was localized to the mitochondrion, and the data obtained by digitonin permeabilization, Western blot analysis, and immunofluorescence indicate that putG3PDH is located within the mitochondrion.
authors.affiliation: Department of Biochemistry, Comenius University, Bratislava, Slovakia.
authors.forename: Ingrid
authors.initials: I
authors.lastname: Škodová
authors.forename: Zdeněk
authors.initials: Z
authors.lastname: Verner
authors.forename: Fréderic
authors.initials: F
authors.lastname: Bringaud
authors.forename: Peter
authors.initials: P
authors.lastname: Fabian
authors.forename: Julius
authors.initials: J
authors.lastname: Lukeš
authors.forename: Anton
authors.initials: A
authors.lastname: Horváth
mesh_terms: D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism*; D005993:Glycerolphosphate Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008928:Mitochondria / Q000201:enzymology* / Q000235:genetics; D021381:Protein Transport; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D014346:Trypanosoma brucei brucei / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000502:physiology; D014353:Trypanosomiasis, African / Q000469:parasitology*
chemical_list: D015800:Protozoan Proteins; D005182:Flavin-Adenine Dinucleotide; D005993:Glycerolphosphate Dehydrogenase