The reversibility of skeletal muscle pyruvate kinase and an assessment of its capacity to support glyconeogenesis.
The kinetics of pyruvate phosphorylation by rabbit skeletal muscle pyruvate kinase (EC 2.7.1.40) has been studied with a coupled assay using P-enolpyruvate carboxylase (EC 4.1.1.31) and malate dehydrogenase (EC 1.1.1.37). The reaction sequence is (See journal for formula). Although the equilibrium of the pyruvate kinase reaction by itself strongly favors pyruvate production, the over-all equilibrium of this coupled system favors the depletion of pyruvate, thus greatly reducing the problem of back reaction during the assay. In addition, the oxidation of NADH by malate dehydrogenase makes it possible to monitor the system with a spectrophotometer. The Michaelis constant of pyruvate kinase was found to be 0.9 mM for ATP and 7 mM for pyruvate, values that agree reasonably well with earlier studies using direct assays. However, the maximum velocity is about 6 mumol of pyruvate phosphorylated/min/mg of enzyme, which is very much faster than that indicated by earlier studies. These results suggest that the metabolic significance of the reverse reaction of muscle pyruvate kinase may have been underestimated. In particular, the data given here suggest that its rate in vivo is probably comparable to the observed rate of glycogen synthesis from lactate, making possible glyconeogenesis in muscle by pyruvate kinase reversal without the need for an enzymatic bypass of the kind employed by liver and kidney.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002262:Carboxy-Lyases / Q000378:metabolism; D006003:Glycogen / Q000096:biosynthesis*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008291:Malate Dehydrogenase / Q000378:metabolism; D009132:Muscles / Q000201:enzymology*; D009243:NAD; D010728:Phosphoenolpyruvate; D010944:Plants; D011188:Potassium / Q000494:pharmacology; D011770:Pyruvate Kinase / Q000378:metabolism*; D011773:Pyruvates; D011817:Rabbits
D011773:Pyruvates; D009243:NAD; D010728:Phosphoenolpyruvate; D000255:Adenosine Triphosphate; D006003:Glycogen; D008291:Malate Dehydrogenase; D011770:Pyruvate Kinase; D002262:Carboxy-Lyases; D008274:Magnesium; D011188:Potassium