Lysosomal phospholipases A1 and A2 of bovine adrenal medulla

Abstract

1. [³²P]Lecithin and [³²P]phosphatidylethanolamine were prepared by incubating rat liver mince with [³²P]phosphate. With these ³²P-labelled phospholipids conditions for the quantitative assay of phospholipase A activity were established. 2. The distribution of phospholipase A activity between subcellular fractions of the bovine adrenal medulla was determined. Phospholipases A₁ and A₂, with pH optima at 4·2 and 6·5 respectively, were found in the large-granule fraction. By means of sucrose-density-gradient centrifugation it was shown that both these phospholipases were localized in lysosomes. 3. Lysosomal phospholipase A₁ catalysed the hydrolysis of [³²P]lecithin and [³²P]phosphatidylethanolamine at the same rate. The enzymic activity was inhibited by 70% in the presence of 10mm-calcium chloride. 4. Lysosomal phospholipase A₂ catalysed the hydrolysis of [³²P]phosphatidylethanolamine more rapidly than it hydrolysed [³²P]lecithin. The hydrolysis of [³²P]phosphatidylethanolamine, but not that of [³²P]lecithin, by phospholipase A₂ was activated by 0·8mm-calcium chloride. However, the hydrolysis of both substrates was inhibited by 8mm-calcium chloride. 5. The significance of the presence of phospholipase activity in lysosomes is discussed in relation to the functions of lysosomes in general and in the adrenal medulla.