A new interpretation of sulfate activation of rabbit muscle glycogen phosphorylase.

It is widely known that sulfate ion at high concentration serves like an allosteric activator of glycogen phosphorylase (GP). Based on the crystallographic studies on GP, it has been assumed that the sulfate ion is bound close to the phosphorylatable Ser14 site of nonactivated GP, causing a conformational change to catalytically-active GP. However, there are also reports that sulfate ion inhibits allosterically-activated GP by preventing the phosphate substrate from attaching to the catalytic site. In the present study, using a high concentration of sulfate ion, significant enhancement of GP activity was observed when macromolecular glycogen was used as substrate but not when smaller maltohexaose was used. In glycogen solution, nonreducing-end glucose residues are localized on the surface of glycogen and are not distributed homogenously in the solution. Using cyclodextrin-immobilized column chromatography, we found that sulfate at high concentration promoted GP-dextrin binding through the dextrin-binding site (DBS) located away from the catalytic site. This result is consistent with the properties of the DBSs found in glycogen-debranching enzyme and β-amylase. Therefore, we propose a new interpretation of the sulfate activation of GP, wherein sulfate ions at high concentration promote glycogen-binding to the DBS directly, and glycogen-binding to the catalytic site indirectly. Our findings were successfully applied to the affinity purification of porcine brain GP.