Skeletal muscle glycogen phosphorylase α kinetics:: Effects of adenine nucleotides and caffeine

This study aimed to determine physiologically relevant kinetic and allosteric effects of Pi, AMP, ADP, and caffeine on isolated skeletal muscle glycogen phosphorylase a (Phos a). In the absence of effectors, Phos a had V-max = 221 +/- 2 U/mg and K-m = 5.6 +/- 0.3 mM P-i at 30 degreesC. AMP and ADP each increased Phos a V-max and decreased K-m in a dose-dependent manner. AMP was more effective than ADP (e.g., 1 muM AMP vs. ADP: V-max = 354 +/- 2 vs. 209 +/- 8 U/mg, and K-m = 2.3 +/- 0.1 vs. 4.1 +/- 0.3 mM). Both nucleotides were relatively more effective at lower P-i levels. Experiments simulating a range of contraction (exercise) conditions in which P-i, AMP, and ADP were used at appropriate physiological concentrations demonstrated that each agent singly and in combination influences Phos a activity. Caffeine (50-100 muM) inhibited Phos a (K-m similar to8-14 mM, similar to 40-50% reduction in activity at 2-10 mM P-i). The present in vitro data support a possible contribution of substrate (P-i) and allosteric effects to Phos a regulation in many physiological states, independent of covalent modulation of the percentage of total Phos in the Phos a form and suggest that caffeine inhibition of Phos a activity may contribute to the glycogen-sparing effect of caffeine.