Rabbit skeletal muscle glycogen synthase. I. Relationship between phosphorylation state and kinetic properties

P. J. Roach, Y. Takeda, J. Larner

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Abstract

Nine samples of purified rabbit skeletal muscle glycogen synthase (UDP glucose:glycogen 4α glucosyltransferase, EC 2.4.1.11) were obtained with alkali labile phosphate contents ranging from 0.27 to 3.49 residues per 85,000 molecular weight subunit. The enzyme samples appeared essentially homogeneous when analyzed by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and had relatively constant specific activity under standard conditions with saturating UDP glucose and glucose 6P concentrations (37.1±1.0 μmol of glucose incorporated/mg/min). When the UDP glucose concentration was varied, deviations from Michaelis Menten kinetics were observed for all samples (Hill slopes of 0.79±0.02), but these deviations were virtually abolished by the presence of 5 mM glucose 6P. Glucose 6P decreased the S(0.5) (concentration required for half maximal rate) for UDP glucose. The plots of activity increase caused by glucose 6P versus glucose 6P concentration became progressively more sigmoid in shape with enzyme samples of higher phosphate content. Both the S(0.5) for UDP glucose and the M(0.5) for glucose 6P (concentration for half maximal activation) were sensitive functions of the alkali labile phosphate content of glycogen synthase. The M(0.5) increased from 3.3 μM at 0.27 phosphate/subunit to 2.7 mM at 3.5 phosphates/subunit, and the S(0.5) varied from 0.75 mM (0.27 phosphate/subunit) to at least 61 mM (2.3 phosphates/subunit). Both parameters increased continuously with phosphate content, with the greatest absolute changes occurring at values greater than 2 residues of phosphates/subunit. The effects of both phosphorylation and activation by glucose 6P appeared to be mediated primarily through alteration of the apparent affinity for UDP glucose. With the phosphorylating conditions used, the results suggested that phosphate could not be introduced into the glycogen synthase molecule without altering its kinetic properties. Further, either the different phosphorylation sites were not equivalent, or else identical sites interacted in determining the kinetic properties of glycogen synthase.

Original languageEnglish (US)
Pages (from-to)1913-1919
Number of pages7
JournalJournal of Biological Chemistry
Volume251
Issue number7
StatePublished - Dec 1 1976

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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