Two Routes for Synthesis of Phosp~oenolp~~ate from C4-dicarboxylic Aicds in Eschkrichia Coli

TWO ROUTES FOR SYNTHESIS OF PHOSP~OENOLP~~ATE FROM C4-DICARBOXYLIC AICDS IN ESCHKRICHIA COLI Eric J. Hansen and Elliot Juni Department of Microbiology The University of Michigan Ann Arbor, Michigan 48104 Received June 28,1974 SUMMARY Mutants of s. coli defective in both phosphoenolpyruvate carboxykinase and phosphoenolpyruvate synthetase are unable to use C4-dicarboxylic acids such as suecinate and malate as carbon and energy sources for growth. Revertants that have restored function for either one of these enzymes can grow in a malate-mineral medium, but at a reduced rate compared with the growth of wildtype cells. g. coli appears to use two pathways for synthesis of phosphoenolpyruvate from C4-dicarboxylic acids. One oi these involves decarboxylation of oxalacetate catalyzed by phosphoenolpyruvate carboxykinase. The second pathway makes use of the combined action of malic enzyme and phosphoenolpyruvate synthetase. INTRODUCTION When microorganisms are grown with a C4 -dicarboxylic acid such as succinate, furmarate, or malate as the sole carbon source they must convert this acid to phosphoenolpyruvate (PEP), which is required for gluconeogenesis as well as for synthesis of pentoses and certain amino acids (1). Phosphoenolpyruvate carboxykinase (PEPCK), which readily decarboxylates oxalacetate to PEP in the presence of a suitable nucleoside triphosphate (2), is generally considered to be the enzyme responsible for net synthesis of PEP during growth with C4-dicarboxylic acid carbon sources (1, 3, 4). A mutant of E. coli lacking a functional PEPCK was reported to be able to grow at slow rates on C4-dicarboxylic acids (3). Another mutant of g. coli was described as being deficient in PEPCK activity and unable to grow with any of the tricarboxylic acid cycle intermediates as a carbon source (4), but this strain was subsequently shown to possess at least two mutations which affect C4 -dicarboxylic acid utilization but not PEPCK activity (5). It is the purpose of the present communication to demonstrate that in addition to the PEPCK route to PEP formation from C4-dicarboxylic