title: Ca2+-independent binding of [3H]phorbol dibutyrate to protein kinase C is supported by protamine and other polycations.
abstract: The activity of the Ca2+- and phospholipid-dependent protein kinase, protein kinase C (PKC), can be modulated by diacylglycerols and phorbol esters. The association of these agents with PKC is, in turn, generally understood to be dependent on Ca2+ and phospholipids. Certain substrates, e.g. protamine sulphate, are known to undergo cofactor-independent phosphorylation by PKC. We report here that, in the presence of such substrates, PKC bound 1,2-dihexanoylglycerol and phorbol dibutyrate in a Ca2+-independent manner. Histone IIIs, which is phosphorylated by PKC only in the presence of Ca2+ and phospholipid, also supported Ca2+-independent binding of 1,2-dihexanoylglycerol and phorbol dibutyrate to PKC, but to a lesser extent than did protamine. Support for Ca2+-independent binding was also exhibited by non-peptide polycations (e.g. DEAE-cellulose DE52), indicating that recognition of the catalytic site is not a prerequisite for this effect. The natural polyamines spermine and putrescine did not have this property, however. The affinity of PKC for phorbol dibutyrate and 1,2-dihexanoylglycerol was found to be unchanged by the presence of substrates or DE52. It is proposed that, in the absence of Ca2+, certain polycations favour expression of the diacylglycerol/phorbol ester binding site by stabilizing the active conformation of PKC.
authors.affiliation: Department of Biochemical Sciences, Wellcome Research Laboratories, Kent, U.K.
authors.forename: N T
authors.initials: NT
authors.lastname: Thompson
authors.forename: R W
authors.initials: RW
authors.lastname: Bonser
authors.forename: H F
authors.initials: HF
authors.lastname: Hodson
authors.forename: L G
authors.initials: LG
authors.lastname: Garland
mesh_terms: D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology*; D002412:Cations / Q000494:pharmacology; D004075:Diglycerides / Q000378:metabolism; D008274:Magnesium / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000378:metabolism*; D010718:Phosphatidylserines / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism; D011073:Polyamines; D000071228:Polyelectrolytes; D011108:Polymers / Q000494:pharmacology; D011479:Protamines / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D002118:Calcium; D002412:Cations; D004075:Diglycerides; D008274:Magnesium; D015240:Phorbol 12,13-Dibutyrate; D010718:Phosphatidylserines; D010743:Phospholipids; D011073:Polyamines; D000071228:Polyelectrolytes; D011108:Polymers; D011479:Protamines; D011493:Protein Kinase C