Inhibition of phosphoenolpyruvate carboxylase from C4 plants by malate and aspartate

Abstract

The activity of phosphoenolpyruvate carboxylase in leaf extracts of C4 plants has been shown to be inhibited by the C4 acids malate and aspartate. The magnitude of inhibition observed with phosphoenolpyruvate carboxylase from Digitaria sanguinalis leaves was dependent on the pH of the assay mixture and the concentration of phosphoenolpyruvate (PEP), Mg2+. and the C4 acid inhibitor. The percentage inhibition decreased with increasing pH up to 8.5. At rate-limiting concentrations of phosphoenolpyruvate the pH optimum of the enzyme was 7.8, whereas the enzyme partially inhibited by malate or aspartate had a pH optimum of roughly 8.3. Malate and aspartate inhibited the enzyme by increasing the apparent Km for phosphoenolpyruvate without altering the maximum velocity, such that inhibition was greatest at rate-limiting concentrations of PEP but was not observed at saturating (3 mM) substrate levels. Glucose-6-P. an activator of PEP carboxylase, decreased the apparent Km for PEP and reversed the inhibition by malate and aspartate. Inhibition was also dependent on the concentration of Mg2+. At 1 mM Mg2+, 1 mM malate caused 60% inhibition, but at 5 mM Mg2+ the inhibition was reduced to 15%. Mg2+ at concentrations from 1 to 5 mM had no effect on the apparent Km for PEP. With Sephadex extracts of PEP carboxylase from leaves of D. sanguinalis, malate and aspartate were equally inhibitory, with roughly 2 mM levels required for 50% inhibition. Both malate and aspartate were inhibitory with PEP carboxylase extracted from leaves of a variety of C4∙ but not C3∙ plants. The significance of the observed inhibition and activation of PEP carboxylase by metabolites is discussed in relation to C4 photosynthesis.