Rapid diglyceride phosphorylation in isolated bovine rod outer segments

Abstract

When isolated bovine rod outer segment fragments were incubated with [γ- 32P]ATP, 32P, as revealed by autoradiography, was rapidly incorporated into rhodopsin bands on sodium dodecyl sulfate polyacrylamide gels, and into a low M r lipid band. Incorporation of 32P into rhodopsin was light-dependent, but labeling of the lipid band was not. A single phosphorylated product, phosphatidic acid, was identified by 2-dimensional thin layer chromatography and by high pressure liquid chromatography of the corresponding glycerophosphate ester. Incorporation of label into phosphatidic acid was detected as early as 15 sec following start of incubation and the product was stable for at least 30 min. No other products were detected, indicating that under the experimental conditions phosphatidic acid was not metabolized to other phospholipids. Up to 1 mol phosphatidic acid was formed per 18 to 40 mol rhodopsin present.