STRUCTURAL-CHANGES IN GLYCOGEN-PHOSPHORYLASE AS REVEALED BY CROSS-LINKING WITH BIFUNCTIONAL DIIMIDATES - PHOSPHORYLASE-B

Glycogen phosphorylase b was cross-linked with a homologous series of diimidates (maximal effective lengths, 3.7-14.5 Å) in the presence of various activators, inhibitors, and substrates under conditions where no polymers were formed. The cross-link products were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the gels were evaluated by densitometry. From the changes of cross-link patterns the following main conclusions were drawn. (1) In dimeric phosphorylase b there are at least two lysyl pairs at the subunit interface (contact m) whose ε-NH2 groups are within 3.7 Å. (2) When dimeric phosphorylase b associates to form tetramers, the nearest two lysyl ε-NH2 groups are ~8 Å apart at the interdimer interface (contact d). (3) The conformational change caused by adenosine 5'-monophosphate (AMP) promotes cross-linking at contact m and induces tetramerization of the enzyme with consequential cross-linkability at contact d. (4) In contrast to AMP, the other activator, inosine monophosphate, does not elicit any change in cross-linking. (5) Adenosine 5'-diphosphate (ADP) and adenosine 5'-triphosphate (ATP) diminish cross-link formation between NH2 groups located within a distance of 4-9 Å at contact m. (6) Glucose 1-phosphate mimicks the effect of ADP, but when applied together with AMP it amplifies the effect of the latter (heterotropic interaction). (7) Glucose, caffeine, and glycogen decrease cross-link formation with the longer reagents owing to the dissociation of tetrameric phosphorylase b. The binding of glucose 6-phosphate does not seem to induce structural changes detectable in the cross-linkability of contact m. All four effectors diminish the influence of AMP. © 1979, American Chemical Society. All rights reserved.