Phosphoenolpyruvate carboxylase activity from Avena coleoptile tissue. Regulation by H+ and malate

Abstract

Preparations of phosphoenolpyruvate (PEP) carboxylase activity from Avenu sativa coleoptile tissue were assayed by measuring the incorporation of labelled bicarbonate into a derivative of oxaloacetic acid or by coupling oxaloacetic acid production to malate dehydrogenase activity and the oxidation of NADH. Malate inhibition of PEP carboxylase activity was found to be noncompetitive, was not due to a mass action effect on the coupled enzyme system or to chelation of Mg²⁺, and probably involved direct inhibition of the enzyme by malate. Maximal PEP carboxylase activity was exhibited around pH 8.0 and increased 125% between pH 7.0 and pH 7.6. Inhibition by 4 mML-malate was virtually complete at pH 7.0 and decreased to 10% inhibition at pH 8. This information is discussed in the light of data which demonstrates that in response to IAA. coleoptile tissue accumulates malate and secretes H⁺. The regulatory properties of PEP carboxylase are consistent with a role in malate production which could resist increases in intracellular pH resulting from an auxin-stimulated H⁺ efflux.