Adenosine 5'-phosphosulfate (APS) kinase: diagnosing the mechanism of substrate inhibition.
Adenosine 5'-phosphosulfate (APS) kinase is subject to strong substrate inhibition by APS. The inhibition has been variously reported to be uncompetitive with respect to MgATP (resulting from the formation of a dead-end E. APS. MgADP complex) or competitive with MgATP (resulting from the formation of a dead-end E. APS complex). It is shown that these two types of substrate inhibition can be differentiated for ordered kinetic mechanisms by simple inspection of the v versus [APS] plots at different fixed concentrations of MgATP. Linear diagnostic plots are unnecessary. One diagnostic feature is the changing position of [APS]opt, the concentration of APS that yields the peak velocity. In the uncompetitive system, [APS]opt decreases asymptotically to a limit as the fixed [MgATP] is increased, while in the competitive system, [APS]opt increases continuously as the fixed [MgATP] is increased. A second (and more easily discerned) diagnostic feature is that, at any given inhibitory level of APS, enzyme activity relative to the velocity at [APS]opt (v/vopt) decreases as the fixed [MgATP] is increased in the uncompetitive system, while in the competitive system the relative activity increases as the fixed [MgATP] is increased. Normalized plots of v/vopt versus [APS] clearly display these distinguishing characteristics. The method confirmed that Penicillium chrysogenum APS kinase exhibits uncompetitive inhibition by APS.
MacRae
I J
IJ
Section of Molecular and Cellular Biology, University of California, One Shields Avenue, Davis, California, 95616, USA.
Segel
I H
IH
D001667:Binding, Competitive; D004789:Enzyme Activation; D007700:Kinetics; D008432:Mathematical Computing; D008954:Models, Biological; D010408:Penicillium chrysogenum / Q000201:enzymology; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D013379:Substrate Specificity
D017853:Phosphotransferases (Alcohol Group Acceptor); C019617:adenylylsulfate kinase