Regulation of phospholipase D activity by neutral lipids in egg-yolk phosphatidylcholine small unilamellar vesicles and by calcium ion in aqueous medium.

Hydrolysis activity of phospholipase D from Streptomyces chromofuscus (PLD) was studied in small unilamellar vesicles (SUV) of egg yolk phosphatidylcholine (PC). The enzyme was associated with PC-SUV in a Ca(2+)-dependent manner. Both apparent maximum velocity, Vmax(app), and reciprocal of apparent Michaelis constant, i.e., apparent binding constant, 1/Km(app), increased with Ca2+ concentration, and the maximum values of these kinetic parameters were obtained at about 20 microM Ca2+. Incorporation of 1,2-diacylglycerol (DAG), cholesterol (Chol) or alpha-tocopherol (Toc) into PC-SUV induced shift of the antisymmetric PO2- stretching band of PC to lower frequency. The neutral lipids in SUV brought about increase of the Vmax(app) value (Yamamoto et al. (1993) Biochim. Biophys. Acta 1145, 293-297). On the basis of these findings we discussed the regulation of PLD activity in terms of the Ca(2+)-dependent complex formation of PLD with SUV, and the enhancement of susceptibility of the P-O bond in PC molecule by neutral lipids.