Studies on the Allosteric Effectors and Some Properties of Phosphoenolpyruvate Carboxylase from Escherichia coli *

Abstract

With partially purified phosphoenolpyruvate (PEP) carboxylase [EC 4.1.1. 31] of Escherichia coli W, the following observations were made which suggest the presence of an allosteric regulatory site on the enzyme for binding with L-aspartate, an inhibitor of the enzyme: (i) The enzyme was specifically protected by aspartate against heat inactivation. (ii) Oxaloacetate, the reaction product, showed only a slight inhibition, suggesting that aspartate exerts its action not as an analogue of oxaloacetate. (iii) The sensitivity of the enzyme to aspartate was shown not to be inherent in all PEP carboxylases from the fact that the enzymes from other organisms such as spinach leaves and Thiobacillns thiooxidans were not inhibited by aspartate. Activation by fructose 1, 6-diphosphate (FDP) was confirmed with the E. coli enzyme and the effect of FDP on the enzyme affinity for PEP and on the inhibition by aspartate was studied. Activation by cyclohexylammonium ions were confirmed and discussion was made on the “pseudo-cooperativity of PEP” which was observed when cyclohexylammonium PEP was employed as substrate. Strong inhibition by chloride ions was found and the effect of the ions on the activation by CoASAc was studied.