Carbon dioxide metabolism by Capnocytophaga ochracea: identification, characterization, and regulation of a phosphoenolpyruvate carboxykinase.
Cell suspensions of Capnocytophaga ochracea incorporated [14C]NaHCO3 into a major four-carbon fermentation product, succinate, and cell-free extracts from this organism contained high levels of phosphoenolpyruvate carboxykinase (PEPCK). PEPCK is the major, if not the only, CO2(HCO-3)-fixing enzyme in C. ochracea since cell-free extracts were devoid of pyruvate-dependent and other phosphoenolpyruvate (PEP)-dependent CO2(HCO-3)-fixing enzymes. The reaction products of the enzyme, which was partially purified by diethylaminoethylcellulose column chromatography, were identified as adenosine 5'-triphosphate (ATP) and oxalacetate. The enzyme showed maximum activity when manganese (Mn2+) was the divalent cation in the incubation mixture, and it had an absolute requirement for the nucleoside 5-'diphosphate adenosine 5'-diphosphate (ADP). PEPCK showed a sigmoidal kinetic response to the Mn2+ concentration and homotropic interactions in its kinetic responses to each of its three substrates PEP, ADP, and CO2(HCO-3). The (S)0.5v values for Mn2+, PEP, ADP, and CO2(CHO-3) were approximately 0.08, 0.3, 0.1, and 10 mM, respectively, and Hill coefficients for these same ligands were 2.60, 1.7, 1.9, and 3.0, respectively. In addition, C. ochracea PEPCK is under metabolic control by the nucleoside -5'-triphosphate ATP, and it also showed a sigmoidal kinetic response to this allosteric effector. The Hill coefficient for ATP was 2.70.
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology; D001639:Bicarbonates / Q000378:metabolism; D002245:Carbon Dioxide / Q000378:metabolism; D002413:Cations, Divalent / Q000494:pharmacology; D041964:Cytophagaceae / Q000201:enzymology*; D005947:Glucose / Q000378:metabolism; D007700:Kinetics; D010728:Phosphoenolpyruvate / Q000378:metabolism; D010729:Phosphoenolpyruvate Carboxykinase (GTP) / Q000302:isolation & purification / Q000378:metabolism*; D013386:Succinates / Q000096:biosynthesis
D001639:Bicarbonates; D002413:Cations, Divalent; D013386:Succinates; D002245:Carbon Dioxide; D000244:Adenosine Diphosphate; D010728:Phosphoenolpyruvate; D000255:Adenosine Triphosphate; D010729:Phosphoenolpyruvate Carboxykinase (GTP); D005947:Glucose