A study of phosphoglycerate kinase in human erythrocytes. II. Kinetic properties.
Kinetic studies on phosphoglycerate kinase (EC 2.7.2.3) were performed in the forward reaction leading from 1,3-diphosphoglycerate to 3-phosphoglycerate. Substrate activation was observed at fixed levels of ADP or Mg2+ and varying concentrations of 1,3-diphosphoglycerate. A biphasic curve was obtained in both linear and double reciprocal plots demonstrating two Km values (Km1 1.9 - 10(-6) and Km2 9.8 - 10(6) M). Michaelis-Menten-type kinetics were observed in both the linear and double reciprocal plots at fixed levels of 1,3-diphosphoglycerate of ADP and varying concentrations of Mg2+. Apparent Michaelis-Menten kinetics were observed in linear plots when conditions of fixed concentrations of 1,3-diphosphoglycerate or Mg2+ were maintained with varying concentrations of ADP. However, the double-reciprocal plots demonstrated biphasic curves with two Km values (Km1 1.7 - 10(-5) and Km2 1.0 - 10(-4)M). Apparent negative cooperativity was observed with respect to 1,3-diphosphoglycerate and ADP. Phosphoglycerate kinase activity was found to be inhibited by AMP and 2,3-diphosphoglycerate. Substrate activation by 1,3-diphosphoglycerate was maintained in the presence of AMP or 2,3-diphosphoglycerate but at a reduced level of enzyme activity. AMP was found to inhibit enzyme activity non-competitively with respect to 1,3-diphosphoglycerate, ADP and Mg2+. 2,3-Diphosphoglycerate inhibits phosphoglycerate kinase activity with respect to 1,3-diphosphoglycerate, ADP and Mg2+. 2,3-Diphosphoglycerate inhibits phosphoglycerate kinase activity non-competitively with respect of 1,3-diphosphoglycerate.
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000249:Adenosine Monophosphate / Q000494:pharmacology; D001667:Binding, Competitive; D004163:Diphosphoglyceric Acids / Q000378:metabolism; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D010735:Phosphoglycerate Kinase / Q000097:blood*
D004163:Diphosphoglyceric Acids; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; D010735:Phosphoglycerate Kinase; D008274:Magnesium