Kinetics of Malic Dehydrogenase Inhibition by 2,4-Dichlorophenoxyacetic Acid

In the course of an investigation of the uncoupling effect of 2,4-D on phosphorylation in plant mitochondria (13), it became evident that this compound exerted two effects on the balance between oxidation of certain of the TCA cycle substrates and associated phosphorylations. In no case was it possible to observe a stimulation of the rate of oxidatioIn concomitant with the inhibition of phosphate esterification, but with malate and citrate a substantial inhibition of oxygen uptake by mitochondria was caused by high concentrations of 2,4-D. This inhibition was lacking when mitochondria were oxidizing succinate, a-ketoglutarate or DPNH. These facts appeared to point toward an interference by 2,4-D with the DPN-requiring dehydrogenases of the miitochondria. It was additionally apparent that this effect on oxidation was independent of uncoupling by 2,4-D, since it occurred at a higher concentration than that causinig a reduction in P/O ratios, the uncoupling effect wvas manifested with substrates whose oxidation was not inhibited, and the inhibitory effect was apparently exerted upon thie first electron transfer step in the oxidation of susceptible substrates, i.e.. preceding the oxidation of re(luced pyridine nucleotide, and thus before the first phosphorylation reaction. The inhibition of respiration in intact cells by 2,4-D has been frequently reported (3), and some years previously an inhibition of the activity of malic dehydrogenase and similar enzymes by naphthalene acetic acid in cell-free preparations or partially purified enzyme preparations was noted (2). The work reported here represents a quantitative investigation of the inhibition by 2,4-D of the activity of malic dehydrogenase isolated from red beet roots and presents kinetic evidence regarding the mechanism by which this inhibition occurs.