Cloning of a phosphatidic acid-preferring phospholipase A1 from bovine testis

We report the molecular cloning and expression of a phosphatidic acid-preferring phospholipase A(1) from bovine testis, The open reading frame encoded am 875-amino acid protein with a calculated molecular mass of 97,576 daltons and a pI of 5.61. The sequence included a region similar to a lipase consensus sequence containing the putative active site serine and also included a potential, coiled-coil-forming region. Expression of the open reading frame in COS1 cells resulted in a 28-44-fold increase in phosphatidic acid phospholipase A(1) activity over that of control cells, Mutation of the putative active site serine (amino acid 540) demonstrated that it was essential far this increase in enzyme activity. Northern blot analysis revealed at. least five different messages with the highest overall message levels in mature testis, but detectable message in all tissues examined. Two possible alternately spliced regions in the spent reading frame also were identified. Finally, a search of the data base identified six related proteins: a potential counterpart of the phospholipase A(1) in Caenorhabditis elegans, two putative lipases in yeast, and three proteins separately encoded by the drosophila retinal degeneration B gene and its mouse and human homologues.