Publications

• Characterization of a membrane-bound biotin-containing enzyme: oxaloacetate decarboxylase from Klebsiella aerogenes.

  Dimroth P.

  Oxaloacetate decarboxylase from Klebsiella aerogenes is firmly bound to the cytoplasmic membrane, from which it can be solubilized with nonionic detergents. The solubilized enzyme behaved like the membrane-bound enzyme with respect to its inhibition by avidin and to the requirement of sodium ions ... >> More

  Oxaloacetate decarboxylase from Klebsiella aerogenes is firmly bound to the cytoplasmic membrane, from which it can be solubilized with nonionic detergents. The solubilized enzyme behaved like the membrane-bound enzyme with respect to its inhibition by avidin and to the requirement of sodium ions for catalytic activity. The decarboxylase was purified 4.5-fold over the solubilized membrane extract by conventional means. Dodecyl-sulfate disc-gel electrophoretic analysis indicated that the enzyme consists of polypeptides of a single size. The molecular weight of these polypeptides is 68000. Radioactive biotin was incorporated specifically into these polypeptide chains upon growth of the bacteria in the presence of the radioactive vitamin. Biotin as the prosthetic group of oxaloacetate decarboxylase is now firmly established. The enzyme in the absence of detergent occurs in a highly aggregated form which elutes in the exclusion volume of a Biogel A 1.5 m column. The reported inhibition of oxaloacetate decarboxylase by citrate could not be repeated. On the other hand oxalate, 2-oxomalonate and glyoxylate proved to be very potent inhibitors of the decarboxylase. The stereochemical course of the oxaloacetate decarboxylation reaction was determined starting from stereospecifically labelled malates, which by malate dehydrogenase and oxaloacetate decarboxylase were converted to chiral pyruvates. The chirality of these pyruvates was analysed via their conversion to acetates and malates by determining the extent of tritium retention upon incubation of the latter with fumarase. It was found that oxaloacetate decarboxylation occurs stereospecifically with retention of configuration. << Less

  Eur J Biochem 115:353-358(1981) [PubMed] [EuropePMC]

• The role of biotin and sodium in the decarboxylation of oxaloacetate by the membrane-bound oxaloacetate decarboxylase from Klebsiella aerogenes.

  Dimroth P.

  The biotin-containing oxaloacetate decarboxylase from Klebsiella aerogenes catalyzed the Na+-dependent decarboxylation of oxaloacetate to pyruvate and bicarbonate (or CO2) but not the reversal of this reaction, not even in the presence of an oxaloacetate trapping system. The enzyme catalyzed an av ... >> More

  The biotin-containing oxaloacetate decarboxylase from Klebsiella aerogenes catalyzed the Na+-dependent decarboxylation of oxaloacetate to pyruvate and bicarbonate (or CO2) but not the reversal of this reaction, not even in the presence of an oxaloacetate trapping system. The enzyme catalyzed an avidin-sensitive isotopic exchange between [1-14C]pyruvate and oxaloacetate, which indicated the intermediate formation of a carboxybiotin enzyme. Sodium ions were not required for this partial reaction, but promoted the second partial reaction, the decarboxylation of the carboxybiotin enzyme, thus accounting for the Na+ requirement of the overall reaction. Therefore, the 14CO2-enzyme which was formed upon incubation of the decarboxylase with [4-15C]oxaloacetate, could only be isolated if Na+ ions were excluded. Preincubation of the decarboxylase with avidin also prevented its labelling with 14CO2. The isolated 14CO2-labelled oxaloacetate decarboxylase revealed the following properties. It was slowly decarboxylated at neutral pH and rapidly upon acidification. The 14CO2 residues of the 14CO2-enzyme could be transferred to pyruvate yielding [4-14C]oxaloacetate. In the presence of Na+ this 14CO2 transfer was repressed by the simultaneous decarboxylation of the 14CO2-enzyme. However, Na+ alone was insufficient as a cofactor for the decarboxylation of the isolated 14CO2-enzyme, since this required pyruvate in addition to Na+. It is therefore concluded that the decarboxylation of oxaloacetate proceeds over a CO2-enzyme--pyruvate complex and that free CO2-enzyme is an abortive reaction intermediate. The activation energy of the enzymic decarboxylation of oxaloacetate changed with temperature and was about 113 kJ below 11 degrees C, 60 kJ between 11 degrees C and 31 degrees C and 36 kJ between 31--45 degrees C. << Less

  Eur J Biochem 121:435-441(1982) [PubMed] [EuropePMC]