3-Methyleneoxindole Reductase of Peas

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Abstract
A 100-fold purification of a reduced triphosphopyridine nucleotide/3-methyleneoxindole reductase of peas has been achieved using conventional protein fractionation procedures. Reduced diphosphopyridine nucleotide is 25-fold less effective than reduced triphosphopyridine nucleotide as the reductant. The preparation is free of other reductase activities including those linking the oxidation of reduced pyridine nucleotide coenzymes to the reduction of cytochrome c; vitamins K1, K2, and K3; O2; nitrate; oxidized glutathione; and thiazolyl blue tetrazolium. The affinity of the enzyme for 3-methyleneoxindole (Ks = 0.5 mm 3-methyleneoxindole) is relatively high. It is, therefore, reasonable to assume that 3-methyleneoxindole is the normal substrate. The enzyme is inhibited by indole-3-acetic acid, indole-3-aldehyde, and by l-naph-thaleneacetic acid. While these are not especially powerful inhibitors (K1 = 1.9-4.0 mm) the competitive relationship with 3-methyleneoxindole indicates that significant inhibition might occur at low intracellular concentrations of the substrate.