Purification of lysosomal phospholipase A. Evidence for multiple isoenzymes in rat liver.

Five isoenzymes of acid phospholipase A have been isolated and purified from the liver of rats.The enzymes were solubilized by freezing and thawing of a composite lysosomal fraction in dilute buffer and purified by chromatography with hydroxyapatite, DEAE-cellulose, concanavalin A-Sepharose, chromatofocusing, and gel filtration.All of the isoenzymes are glycoproteins varying in molecular weight from 90,000 to 15,000.The two major forms have molecular weights of 34,000 and 44,000, respectively, and their approximate PI values are 5.2 and less than 4.0, based on chromatofocusing.They have been purified 17,500-and l,glO-fold, respectively.All of the recovered isoenzymes appear to have intrinsic lysophospholipase activity and are phospholipases of the AI type.The five phospholipases AI did not require Ca2+ and were not inhibited by EDTA or p - bromophenacyl bromide.Triton X-100 and Hg2+ stimulated the activity of the phospholipase AI isoenzymes.The two major isoenzymes degraded all of the major phosphoglyceride classes, but phosphatidylglycerol and cardiolipin were hydrolyzed most rapidly.These five isoenzymes may represent precursor and processed forms of lysosomal phospholipase A similar to those which have been reported for several other lysosomal hydrolases (Hasilik, A., and Neufeld, E. (1980)