We investigated the inhibitory effect of calphobindin I (CPB I) on protein kinase C activity. CPB I inhibited protein kinase C activity in a concentration-dependent manner with an IC50 value of 70 nM. The inhibition of protein kinase C by CPB I was Ca(2+)-dependent, and was partially overcome by increasing the concentration of phosphatidylserine. Since CPB I is widely distributed in various organs and cells, it is presumed that CPB I is an endogenous inhibitor of protein kinase C.