Protein kinase C interaction with calcium: a phospholipid-dependent process.
The calcium-binding properties of calcium- and phospholipid-dependent protein kinase C (PKC) were investigated by equilibrium dialysis in the presence and the absence of phospholipids. Calcium binding to PKC displayed striking and unexpected behavior; the free proteins bound virtually no calcium at intracellular calcium concentrations and bound limited calcium (about 1 mol/mol of PKC) at 200 microM calcium. However, in the presence of membranes containing acidic phospholipids, PKC bound at least eight calcium ions per protein. The presence of 1 microM phorbol dibutyrate (PDBu) in the dialysis buffer had little effect on these calcium-binding properties. Analysis of PKC-calcium binding by gel filtration under equilibrium conditions gave similar results; only membrane-associated PKC bound significant amounts of calcium. Consequently, PKC is a member of what may be a large group of proteins that bind calcium in a phospholipid-dependent manner. The calcium concentrations needed to induce PKC-membrane binding were similar to those needed for calcium binding (about 40 microM calcium at the midpoint). However, the calcium concentration required for PKC-membrane binding was strongly influenced by the phosphatidylserine composition of the membranes. Membranes with higher percentages of phosphatidylserine required lower concentrations of calcium. These properties suggested that the calcium sites may be generated at the interface between PKC and the membrane. Calcium may function as a bridge between PKC and phospholipids. These studies also suggested that calcium-dependent PKC-membrane binding and PKC function could be regulated by a number of factors in addition to calcium levels and diacylglycerol content of the membrane.
Bazzi
M D
MD
Department of Biochemistry, University of Minnesota, St. Paul 55108.
Nelsestuen
G L
GL
D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D003956:Dialysis; D010703:Phorbol Esters / Q000494:pharmacology*; D010743:Phospholipids / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*
D010703:Phorbol Esters; D010743:Phospholipids; D011493:Protein Kinase C; D002118:Calcium