Properties of pyruvate carboxylase of the facultative methylotrope Pseudomonas oleovorans
Loginova, N.V.; Sokolov, A.P.; Trotsenko, I.A.
Biokhimiia 47(10): 1629-1634
1982
ISSN/ISBN: 0320-9725 PMID: 7171645 Document Number: 187895
Pyruvate carboxylase of the facultative methylotroph P. oleovorans was purified 40-fold by ammonium sulfate fractionation, gel filtration on Ultrogel AcA 34, ion-exchange chromatography on DEAE-Biogel A and concentration on DEAE-Sepharose CL-6B. The enzyme exerts its maximal activity in the presence of Mg2+ (pH 7.5, 40.degree. C), is unstable and completely inactivated within 6 h at 25.degree. C. In the presence of Mg2+, monovalent cations stimulate the enzyme activity. The MW of pyruvate carboxylase as determined by gel filtration on Sepharose CL-6B is 300,000. The enzyme molecule contains biotin. The apparent Km values for pyruvate, ATP and HCO30- are 1.77, 0.19 and 0.23 mM, respectively. The enzyme is inhibited by aspartate, malate, oxaloacetate and is activated by citrate, isocitrate and phosphosugars. The role of pyruvate carboxylase in methylotrophic metabolism of P. oleovorans is discussed.