EXTENSIVE CAMP-DEPENDENT AND CAMP-INDEPENDENT PHOSPHORYLATION OF MICROTUBULE-ASSOCIATED PROTEIN-2

Abstract

Microtubule-associated protein 2 (MAP 2) is the major substrate for phosphorylation in purified preparations of [calf] brain microtubules. Phosphorylation is catalyzed by a type II cAMP-dependent protein kinase tightly associated with MAP 2 itself. The extent of MAP 2 phosphorylation wsa examined by its associated protein kinase. Using Pi assay, MAP 2 contained from 8-13 mol of phosphate/mol of protein was isolated. The catalytic subunit of the MAP 2-associated kinase catalyzed the incorporation of additional phosphate to a final level of 20-22 mol/mol of MAP 2. Potato acid phosphatase was used to remove phosphate from MAP 2. Rephosphorylation of acid phosphatase-treated MAP 2 resulted in maximal incorporation of 13 mol of phosphate/mol of MAP 2. The rates and extent of [32P] phosphate incorporation into as isolated and dephosphorylated MAP 2 were identical, and phosphate was incorporated into identical peptides in the 2 preparations. MAP 2 contains as many as 13 cAMP-dependent phosphorylation sites and approximately 8 phosphates of as yet undetermined origin.