Inhibition of protein kinase C by annexin V.
Annexin V is a protein of unknown biological function that undergoes Ca(2+)-dependent binding to phospholipids located on the cytosolic face of the plasma membrane. Preliminary results presented herein suggest that a biological function of annexin V is the inhibition of protein kinase C (PKC). In vitro assays showed that annexin V was a specific high-affinity inhibitor of PKC-mediated phosphorylation of annexin I and myosin light chain kinase substrates, with half-maximal inhibition occurring at approximately 0.4 microM. Annexin V did not inhibit epidermal growth factor receptor/kinase phosphorylation of annexin I or cAMP-dependent protein kinase phosphorylation of the Kemptide peptide substrate. Since annexin V purified from both human placenta and recombinant bacteria inhibited protein kinase C activity, it is not likely that the inhibitor activity was associated with a minor contaminant of the preparations. The following results indicated that the mechanism of inhibition did not involve annexin V sequestration of phospholipid that was required for protein kinase C activation: similar inhibition curves were observed as phospholipid concentration was varied from 0 to 800 micrograms/mL; the extent of inhibition was not significantly affected by the order of addition of phospholipid, substrate, or PKC, and the core domain of annexin I was not a high-affinity inhibitor of PKC even though it had similar Ca2+ and phospholipid binding properties as annexin V. These data indirectly indicate that inhibition occurred by direct interaction between annexin V and PKC. Since the concentration of annexin V in many cell types exceeds the amounts required to achieve PKC inhibition in vitro, it is possible that annexin V inhibits PKC in a biologically significant manner in intact cells.
Schlaepfer
D D
DD
Department of Biological Chemistry, University of California, Irvine 92717.
Jones
J
J
Haigler
H T
HT
D017304:Annexin A5; D017302:Annexins; D002135:Calcium-Binding Proteins / Q000378:metabolism / Q000494:pharmacology*; D000242:Cyclic AMP / Q000494:pharmacology; D006801:Humans; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D009842:Oligopeptides / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism; D010766:Phosphorylation; D010920:Placenta / Q000737:chemistry; D011257:Pregnancy Proteins / Q000494:pharmacology*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism
D017304:Annexin A5; D017302:Annexins; D002135:Calcium-Binding Proteins; D009842:Oligopeptides; D010743:Phospholipids; D011257:Pregnancy Proteins; C037642:kemptide; D000242:Cyclic AMP; D011494:Protein Kinases; D011493:Protein Kinase C; D009219:Myosin-Light-Chain Kinase