Production of 1,2-diacylglycerol in human erythrocyte membranes exposed to low concentrations of calcium ions

A specific increase in the membrane content of 1,2-diacylglycerol occurred when erythorcytes were lysed at 20 °C in media which did not include a chelator of Ca²⁺ and also when Ca²⁺ was added to haemoglobin-free erythrocyte ghosts which had been prepared in the presence of ethyleneglycol-bis-(β-aminoethylether)-N,N′-tetraacetic acid (EGTA). The maximum increase was about 20-fold. The production of 1,2-diacylglycerol appeared to be caused by an endogenous membrane-bound phospholipase C which was half-maximally activated at less than 1 μM Ca²⁺ and which had access to only about 0.6-0.8% of the cells' glycerolipids. This activity was optimal at pH 7.0-7.2 in the presence of 0.1 mM Ca²⁺; under these conditions diacylglycerol production was complete within 5-10 min. Enzyme activity was markedly decreased at low temperatures, and was abolished by heating at 100 °C for 1 min.