PEPTIDE-SUBSTRATES SUITABLE FOR ASSAYING GLYCOGEN-SYNTHASE KINASE-3 IN CRUDE CELL-EXTRACTS

In this study we describe the characterization and use of new peptide substrates for assaying glycogen synthase kinase-3 (GSK-3) which are b ased on the sequence around the single GSK-3 phosphorylation site in t he translation factor eIF2B. The new peptides offer important advantag es over previous substrates, which were based on the sequence around t he multiple GSK-3 phosphorylation sites in glycogen synthase (GS), for the assay of GSK-3 in cell extracts. In particular, decreases in GSK- 3 activity following, e.g., insulin treatment, are partially or comple tely masked when the GS-based peptides are used but are readily measur ed using the new, eIF2B-based, peptides. The new peptides, unlike thos e based on GS, are therefore suitable for the assay of changes in GSK- 3 activity in cell extracts without the need for prior immunoprecipita tion or ion-exchange chromatography. (C) 1997 Academic Press, Inc.