Abstract

Dihydrofolate reductase was isolated from acetone powder extracts of T. equiperdum. Folic acid was not an effective substrate; indeed, it was a weak inhibitor, the concentration required for 50% inhibition being 3 x 10^-5 M. NADPH was a much more efficient hydrogen donor than NADH. Michaelis constants of 4.3 x 10^-6 M for dihydrofolate and 1.2 x 10^-5 M for NADPH were obtained. Trypanosomal dihydrofolate reductase showed a broad peak of maximal activity between pH 6.0 and 7.7, with activity sharply decreasing above and below those values. Activity increased with increasing temperature within the range of 20°-40°, but fell off sharply above 40°. 4-Amino analogs of folic acid are potent inhibitors of trypanosomal dihydrofolate reductase, the concentration for 50% inhibition by methotrexate, for example, being 2 x 10^-10 M. The trypanosomal reductase differs from those of mammalian and bacterial origin with respect to susceptibility to inhibition by a number of 2,4-diaminoheterocyclic compounds.

A substance (or substances) was isolated from homogenates of T. equiperdum which possessed spectrophotofluorometric properties closely resembling those of 2-amino-4-hydroxypteridines and which effectively replaced folic acid as a growth-promoting factor for Lactobacillus casei.