THE CELL-MEMBRANE OF MYCOPLASMA PENETRANS - LIPID-COMPOSITION AND PHOSPHOLIPASE A(1) ACTIVITY

Analysis of Mycoplasma penetrans membrane lipids revealed that, in addition to large amounts of unesterified cholesterol, M. penetrans incorporated exogenous phospholipids, preferentially sphingomyelin, from the growth medium. The major phospholipids synthesized de novo by M. penetrans were phosphatidylglycerol (PG) and diphosphatidylglycerol (DPG). In vivo labeling of PG and DPG by growing the cells with radioactive palmitate or oleate, followed by snake venom phospholipase A(2) treatment, enabled us to assess the positional distribution of fatty acids in these lipids. Saturated fatty acids were found preferentially in position 2 of the glycerol backbone, and not in position 1 as found elsewhere in nature, while unsaturated fatty acids prefer position 1. M. penetrans membranes contain phospholipase activity of the A(1) type, removing a fatty acid from the sn-1 ester bond of phospholipids. The activity was neither stimulated by Ca2+ nor inhibited by EGTA and had a broad pH spectrum. The substrate specificity of the enzyme was investigated with various natural lipids and with a fluorescent analog of the phosphatidylcholine. The enzyme was equally active toward phosphatidylcholine and phosphatidylglycerol, but did not hydrolyze diphosphatidylglycerol. The enzyme did not act on triacylglycerol, diacylglycerol or cholesteryl ester, but low activity was detected toward monoacylglycerol. The enzyme was heat-sensitive and detergent-sensitive, and was almost completely inhibited by p-bromophenacylbromide (50 mu M), but was not affected by SH reagents. This study is the first one reporting phospholipase A(1) activity in Mollicutes. A possible role of this enzyme in forming lipid mediators upon the interaction of M. penetrans cells with eukaryotic cells is suggested.