The Mg2+ requirements of nonactivated and activated rat liver phosphorylase kinase. Inhibition of the activated form by free Mg2+.
Incubation of rat liver phosphorylase kinase in the presence of MgATP results in a time-dependent increase in activity, i.e., activation. Determination of the magnitude of activation depends, in large part, on the relative concentrations of Mg2+ and ATP used in the phosphorylase kinase activity assay, such that as the Mg2+ to ATP ratio increases less activation is detectable. Prior to activation, maximal activity of nonactivated phosphorylase kinase requires a 2-3-fold molar excess of Mg2+ (i.e., free Mg2+) over ATP. MgATP-dependent activation of the enzyme results in an alteration in the free Mg2+ requirement such that the activity of the activated enzyme is sharply inhibited by the free cation. Inhibition by free Mg2+ of the activated enzyme is rapidly reversed by removal of free Mg2+ but is not affected by addition of Ca2+. Both nonactivated and activated forms of the enzyme appear to be inhibited by free ATP4-. The results show that the use of high concentrations of free Mg2+ in the phosphorylase kinase activity assay can blunt or completely obscure changes in enzyme activity following activation of the enzyme.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000502:physiology; D004789:Enzyme Activation / Q000187:drug effects; D008099:Liver / Q000201:enzymology*; D008274:Magnesium / Q000502:physiology*; D010764:Phosphorylase Kinase / Q000378:metabolism*; D011817:Rabbits; D051381:Rats
D000255:Adenosine Triphosphate; D010764:Phosphorylase Kinase; D008274:Magnesium; D002118:Calcium