Studies of Dlalkyl Ether Phospholipids. II. Requirement for a Liquid-Crystalline Substrate for Hydrolysis by Cabbage Leaf Phospholipase D

Abstract

The hydrolysis of 1,2-ditetradecyl-, 1,2-dihexadecyl-, and 1,2-dioctadecyk-sn-glycero-3-phosphoryl-cholines by the soluble phospholipase D (EC 3.14.4) from cabbage leaf has been compared with that of egg lecithin. When a diethyl ether – chloroform mixture was used as activator low hydrolysis rates were observed for the dialkyl ether phospholipids because of their poor solubility in diethyl ether. Under these conditions the compounds did not inhibit the hydrolysis of egg lecithin. The hydrolytic reaction appeared to take place at the solvent–water interface resulting in a gradual denaturation of the enzyme. When sodium dodecyl sulfate was used as activator faster hydrolysis was evident due in part to a reduction in mesomorphic transition temperature (Tm) revealed by differential thermal analysis. Rates of hydrolysis of the homologous dialkyl ether phospholipids were inversely proportional to their Tm values in excess water. Under conditions required for effective hydrolysis of these compounds they inhibited the hydrolysis of egg lecithin. From studies of hydrolysis rates obtained with various combinations of phosphatidylcholine and phosphatidic acid analogues it was concluded that binding of enzyme to phospholipids and their subsequent hydrolysis required both an appropriate surface charge and a gel to liquid-crystalline phase transition in the substrate.