Properties of l-α-glycerophosphate oxidase and its role in the respiration of Trypanosoma rhodesiense

Abstract

Cell-free extracts of Trypanosoma rhodesiense have been found to contain a cytoplasmic L-[alpha]-glycerophosphate dehydrogenase and a particulate hydrogen-transport system whose rate of reaction with L-[alpha]-glycerophosphate was about 20 times that of other possible hydrogen donors such as reduced diphosphopyridine nucleotide or succinate. The latter enzyme system has been termed L-[alpha]-glycerophosphate oxidase, as 1 g.atom of oxygen was utilized in the oxidation of L-[alpha]-glycerophosphate to dihydroxyacetone phosphate. The optimum activity of the oxidase in phosphate buffer and in 2-amino-2-hydroxymethyl-propane-l,3-diol buffer was about pH 7.4 and pH 8.0 respectively. At pH 7.4 the K[image] was calculated to be 1.3 m[image]. The activity of the oxidase was inhibited by certain thiol reagents but not at all by compounds which inhibit systems containing cytochrome pigments. There was no evidence that pyridine nucleotides are involved in the oxidase reaction. Under conditions where the substrate concentration was not a limiting factor over the experimental period, the rate of uptake of oxygen by the oxidase was about 60 and 110% of that found for the intact trypano-some respiring in either glycerol or glucose respectively. A dialysed cell-free suspension of T. rhodesiense utilized 1 g.atom of oxygen/mole of L-[alpha]-glycerophosphate added but this uptake of oxygen could be doubled when oxidized diphosphopyridine nucleotide was also added. It is suggested that the reactions catalysed by the diphosphopyridine nucleotide-linked dehydrogenase and the oxidase are coupled and constitute a hydrogen-transport system for the oxidation of reduced diphosphopyridine nucleotide. The compatability of this suggestion with previous work on the oxidative catabolism of glucose by T. rhodesiense has been discussed.