The lipid binding, regulatory domain of protein kinase C. A 32-kDa fragment contains the calcium- and phosphatidylserine-dependent phorbol diester binding activity.
Trypsinization of rat brain protein kinase C (80 kDa) into 50- and 32-kDa fragments occurred without inhibition of [3H]phorbol dibutyrate ([3H]PDBu) binding activity. The 50-kDa fragment, the catalytic domain (Inoue, M., Kishimoto, A., Takai, Y., and Nishizuka, Y. (1977) J. Biol. Chem. 252, 7610-7616), was further degraded by trypsin, whereas the 32-kDa fragment was resistant. Protein kinase activity and the [3H]PDBu binding activity were completely separated upon gel filtration of a solution containing Triton X-100/phosphatidylserine mixed micelles and trypsinized protein kinase C. Pooled fractions of the [3H]PDBu binding activity contained a 32-kDa fragment exclusively. The binding of [3H]PDBu to this fragment was dependent on calcium and phosphatidylserine and was of high affinity (Kd = 2.8 nM) and of essentially identical specificity to that of native protein kinase C. It is concluded that the 32-kDa fragment represents a lipid binding, regulatory domain of protein kinase C.
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D000818:Animals; D001667:Binding, Competitive; D001921:Brain / Q000201:enzymology; D002118:Calcium / Q000494:pharmacology*; D005260:Female; D007700:Kinetics; D008823:Micelles; D010446:Peptide Fragments / Q000378:metabolism; D015240:Phorbol 12,13-Dibutyrate; D010703:Phorbol Esters / Q000378:metabolism*; D010718:Phosphatidylserines / Q000494:pharmacology*; D011485:Protein Binding; D011493:Protein Kinase C / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D014357:Trypsin
D008823:Micelles; D010446:Peptide Fragments; D010703:Phorbol Esters; D010718:Phosphatidylserines; D015240:Phorbol 12,13-Dibutyrate; D011493:Protein Kinase C; D014357:Trypsin; D002118:Calcium