No abstract

A novel Ca(2+)-independent phospholipase A2 (PLA2) has recently been purified from the murine macrophage-like cell line P388D1 (Ackermann, E. J., Kempner, E. S., and Dennis, E. A. (1994) J. Biol. Chem. 269, 9227-9233). This enzyme is now shown to be inhibited by palmitoyl trifluoromethyl ketone (PACOCF3), arachidonyl trifluoromethyl ketone (AACOCF3), and a bromoenol lactone (BEL). Both PACOCF3 and AACOCF3 were found to inhibit the macrophage PLA2 in a concentration-dependent manner. PACOCF3 was found to be approximately 4-fold more potent than AACOCF3, with IC50 values of 3.8 microM (0.0075 mol fraction) and 15 microM (0.028 mol fraction), respectively. Reaction progress curves in the presence of either inhibitor were found to be linear, and the PACOCF3.PLA2 complex rapidly dissociated upon dilution. BEL was also found to inhibit the macrophage PLA2 in a concentration-dependent manner, with half-maximal inhibition observed at 60 nM after a 5-min preincubation at 40 degrees C. Inhibition was not reversed after extensive dilution of the enzyme into assay buffer. Treatment of the PLA2 with BEL resulted in a linear, time-dependent inactivation of activity, and the rate of this inactivation was diminished in the presence of PACOCF3. In addition, PLA2 treated with [3H]BEL resulted in the covalent labeling of a major band at M(r) 80,000. Inactivation of the PLA2 by 5,5'-dithiobis(2-nitrobenzoic acid) prior to treatment with [3H]BEL resulted in the near complete lack of labeling consistent with covalent irreversible suicide inhibition of the enzyme. The labeling of a M(r) 80,000 band rather than a M(r) 40,000 band upon treatment with [3H]BEL distinguishes the macrophage Ca(2+)-independent PLA2 from a previously identified myocardial Ca(2+)-independent PLA2 and provides strong evidence that the M(r) 80,000 protein is the catalytic subunit.

Many lipids or lipid-derived products generated by phospholipases acting on phospholipids in membranes are implicated as mediators and second messengers in signal transduction. Our current understanding of the primary sequence relationships within the class of extracellular phospholipase A2's and among the numerous forms of the mammalian phosphatidylinositol-specific phospholipase C's is reviewed. New results suggesting roles for these phospholipases as well as other phospholipases such as phospholipase C and D acting on phosphatidlycholine in generating arachidonic acid for eicosanoid biosynthesis, inositol phosphates for Ca2+ mobilization, and diglyceride for protein kinase C activation through receptor-mediated processes, are discussed. In addition, the possible role of phospholipases acting on sphingolipids such as sphinglomyelinase in generating lipid mediators is considered.

Calculations of the size, shape, and hydration of micelles composed of the nonionic surfactant Triton X-100 were performed based on molecular weight and intrinsic viscosity data. Geometrical considerations show that if the hydrophobic core as well as the whole micelle is spherical, then its structure cannot contain the distinct polar and apolar regions that are classically assumed for micelles. On the other hand, ellipsoids of revolution would be consistent with a classical micellar structure and an oblate rather than a prolate ellipsoid would be most consistent with intrinsic viscosity measurements and volume calculations.

Lipids are produced, transported, and recognized by the concerted actions of numerous enzymes, binding proteins, and receptors. A comprehensive analysis of lipid molecules, "lipidomics," in the context of genomics and proteomics is crucial to understanding cellular physiology and pathology; consequently, lipid biology has become a major research target of the postgenomic revolution and systems biology. To facilitate international communication about lipids, a comprehensive classification of lipids with a common platform that is compatible with informatics requirements has been developed to deal with the massive amounts of data that will be generated by our lipid community. As an initial step in this development, we divide lipids into eight categories (fatty acyls, glycerolipids, glycerophospholipids, sphingolipids, sterol lipids, prenol lipids, saccharolipids, and polyketides) containing distinct classes and subclasses of molecules, devise a common manner of representing the chemical structures of individual lipids and their derivatives, and provide a 12 digit identifier for each unique lipid molecule. The lipid classification scheme is chemically based and driven by the distinct hydrophobic and hydrophilic elements that compose the lipid. This structured vocabulary will facilitate the systematization of lipid biology and enable the cataloging of lipids and their properties in a way that is compatible with other macromolecular databases.