Properties of a mutant Escherichia coli phosphoenolpyruvate carboxylase deficient in coregulation by intermediary metabolites

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Abstract
Phosphoenolpyruvate carboxylase of E. coli is activated by 3 different mechanisms: contiguous by acetyl CoA, precursor by fructose 1,6-bisphosphate and compensatory feedback by CDP. Even though each activator can interact independently with the enzyme, synergistic effects are observed with some combinations, namely, fructose 1,6-bisphosphate or CDP (coregulators), with acetyl CoA. A mutant was isolated that has a phosphoenolpyruvate carboxylase which is refractory to activation by fructose 1,6-bisphosphate and CDP. The mutant enzyme was active primarily as the dimer and to lack cooperativity in substrate binding. The binding of acetyl CoA and substrate was essentially the same as that of the wild-type enzyme. The mutant cells grew extremely slowly on glucose alone as the sole C source. The only defect in the mutant appeared to be the inability of this enzyme to be activated by the coregulators. Coregulation by fructose 1,6-bisphosphate or CDP is apparently essential for the activation in vivo of this enzyme.