Zdeněk Verner, Ingrid Skodová, Simona Poláková, Vladislava Durišová-Benkovičová, Anton Horváth, Julius Lukeš
Biology Centre, Institute of Parasitology, Czech Academy of Sciences, České Budějovice (Budweis), Czech Republic.
Parasitology 2013 MarThe respiratory chain of the procyclic stage of Trypanosoma brucei contains the standard complexes I through IV, as well as several alternative enzymes contributing to electron flow. In this work, we studied the function of an alternative NADH : ubiquinone oxidoreductase (NDH2). Depletion of target mRNA was achieved using RNA interference (RNAi). In the non-induced and RNAi-induced cell growth, membrane potential change, alteration in production of reactive oxygen species, overall respiration, enzymatic activities of complexes I, III and/or IV and distribution of NADH : ubiquinone oxidoreductase activities in glycerol gradient fractions were measured. Finally, respiration using different substrates was tested on digitonin-permeabilized cells. The induced RNAi cell line exhibited slower growth, decreased mitochondrial membrane potential and lower sensitivity of respiration to inhibitors. Mitochondrial glycerol-3-phosphate dehydrogenase was the only enzymatic activity that has significantly changed in the interfered cells. This elevation as well as a decrease of respiration using NADH was confirmed on digitonin-permeabilized cells. The data presented here together with previously published findings on complex I led us to propose that NDH2 is the major NADH : ubiquinone oxidoreductase responsible for cytosolic and not for mitochondrial NAD+ regeneration in the mitochondrion of procyclic T. brucei.
Zdeněk Verner, Ingrid Skodová, Simona Poláková, Vladislava Durišová-Benkovičová, Anton Horváth, Julius Lukeš. Alternative NADH dehydrogenase (NDH2): intermembrane-space-facing counterpart of mitochondrial complex I in the procyclic Trypanosoma brucei. Parasitology. 2013 Mar;140(3):328-37
PMID: 23111000
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