Purification and characterization of phosphoenolpyruvate carboxykinase,a catabolic CO2-fixing enzyme, from Anaerobiospirillum succiniciproducens

J Gen Microbiol. 1993 Feb;139(2):223-8. doi: 10.1099/00221287-139-2-223.

Abstract

Phosphoenolpyruvate (PEP) carboxykinase (EC 4.1.1.49) from the obligate anaerobe Anaerobiospirillum succiniciproducens was purified 18-fold. The enzyme was monomeric, with an Mr of 57,000 +/- 2,000. The enzyme was oxygen stable, had a pH optimum of 6.7-7.1, and was stable from pH 5.0 to 9.0. The enzyme displayed Michaelis-Menten kinetics for the substrate PEP and the cosubstrates bicarbonate and ADP with a Km of 0.54 mM, 17 mM and 0.42 mM, respectively. The enzyme required Mn(2+) or Co(2+) in addition to Mg(2+) to exhibit maximum activity. p-Chloromercuribenzoate inhibited activity and phosphoenolpyruvate protected the enzyme against inactivation, suggesting that an essential cysteine may be in the active site.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Adenosine Diphosphate / metabolism
  • Amino Acid Sequence
  • Amino Acids / analysis
  • Bacteroidaceae / enzymology*
  • Bicarbonates / metabolism
  • Carbon Dioxide / metabolism
  • Isoelectric Point
  • Kinetics
  • Metals / pharmacology
  • Molecular Sequence Data
  • Phosphoenolpyruvate / metabolism
  • Phosphoenolpyruvate Carboxykinase (GTP) / chemistry
  • Phosphoenolpyruvate Carboxykinase (GTP) / isolation & purification*
  • Phosphoenolpyruvate Carboxykinase (GTP) / metabolism

Substances

  • Amino Acids
  • Bicarbonates
  • Metals
  • Carbon Dioxide
  • Adenosine Diphosphate
  • Phosphoenolpyruvate
  • Phosphoenolpyruvate Carboxykinase (GTP)