Adenosine 5'-diphosphate as an allosteric effector of phosphorylase kinase from rabbit skeletal muscle.
Equilibrium binding and activity studies indicate that adenosine 5'-diphosphate binds to phosphorylase kinase with high affinity at a site, or sites, distinct from the catalytic site. Equilibrium dialysis at pH 6.8 and 8.2, with and without Mg2+, and with phosphorylated and nonphosphorylated enzyme preparations revealed approximately 8 ADP binding sites per alpha 4 beta 4 gamma 4 delta 4 hexadecamer, with Kd values ranging from 0.26 to 17 microM. Decreasing the pH from 8.2 to 6.8 or removing the Mg2+ enhanced the affinity for ADP. At pH 6.8, ADP stimulated the phosphorylase conversion and autophosphorylation activities of the nonactivated enzyme. Analogs of ADP with modifications at the 2'-, 3'-, and 5'-positions allowed determination of structural requirements for the stimulation of activity. ADP seems to alter the conformation of the beta subunit because addition of the nucleotide inhibits its dephosphorylation by phosphoprotein phosphatase and its chemical cross-linking by 1,5-difluoro-2,4-dinitrobenzene. The binding affinities and effects of ADP suggest that it may function physiologically as an allosteric effector of phosphorylase kinase.
D000244:Adenosine Diphosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000494:Allosteric Regulation; D000495:Allosteric Site / Q000187:drug effects; D000818:Animals; D003432:Cross-Linking Reagents; D004139:Dinitrofluorobenzene / Q000031:analogs & derivatives / Q000494:pharmacology; D046911:Macromolecular Substances; D009132:Muscles / Q000201:enzymology*; D010764:Phosphorylase Kinase / Q000378:metabolism*; D010766:Phosphorylation; D011817:Rabbits; D013329:Structure-Activity Relationship
D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D004139:Dinitrofluorobenzene; D010764:Phosphorylase Kinase; C002041:1,5-difluoro-2,4-dinitrobenzene