[Reaction between adenosine-5'-chloromethylphosphonate and the allosteric center of phosphorylase B].

The interaction between phosphorylase B and an AMP analog, adenosine-5'-chloromethylphosphonate, is found to be irreversible. Their binding stechiometry is calculated from the differential absorption spectrum. Maximal inhibition is reached when 1,3--1,5 moles of the analogue is bound per mole of monomer phosphorylase B. The enzyme-inhibitory complex exhibited 50% activity is characterized by a sigmoid curve of the reaction rate dependency on the substrate concentration, by a decrease of the affinity to glucose-1-phosphate and the maximal rate, and by an increase of Hill's coefficient. Similar SH-groups titration curves were obtained for modified and native phosphorylase in the presence of AMP. Apophosphorylase was incapable of the complete reactivation by pyridoxalphosphate in the presence of adenosine-5'-chloromethylphosphonate. The complex of phosphorylase B and the AMP analogue is found to be inhibited by glucose-6-phosphate, like the native enzyme in the presence of AMP. The results of ultracentrifugation and disc electrophoresis show that the AMP analogue contributes the formation of the tetrameric form of the enzyme. The data obtained indicate the tight binding of adenosine-5'-chloromethylphosphonate in the active site of phosphorylase B. The properties of the complex confirm the importance of the phosphate group for the AMP binding in the allosteric site and for the enzyme activation.