CHARACTERIZATION OF CAMP-DEPENDENT PROTEIN-KINASE FROM THE PUPAL WING EPIDERMIS OF MANDUCA-SEXTA

Cyclic AMP-dependent protein kinase (cAMP-PK) activity in the wing epidermis of day zero pupae of Manduca sexta was characterized. The preferred exogenous substrates were histones, subfractions H1 and H2b, casein and protamine sulfate; histone H2a was only phosphorylated moderately, while free base protamine and bovine serum albumin were poor substrates for cAMP-PK. cAMP-PK activity required Mg2+ and was optimal in the presence of 1 mM Mg2+. Co2+ and Mn2+ did not substitute for Mg2+, and Ca2+ inhibited cAMP-PK activity. The effective concentration of cAMP for activation of the cAMP-PK was substantially lower than that of cGMP (EC50 1.3 × 10-8 and 1.2 × 10-6 M, respectively). The type II isozyme of cAMP-PK comprised approx. 75% of the total cytosolic wing cAMP-PK as determined by DEAE anion exchange chromatography. Photoaffinity labeling of the whole cell homogenate with 8-azido cAMP revealed the presence of only type II isozyme. The distribution of the cAMP-PK isozymes was also determined for whole cell homogenates of brain, prothoracic glands, hemolymph, trachea, nerve cord, fat body, muscle and midgut. © 1990.