Inhibition of the plasma glycosylphosphatidylinositol-specific phospholipase D by synthetic analogs of lipid A and phosphatidic acid.
Glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD), a plasma enzyme with extensive sequence similarity to integrin alpha subunits, is inhibited by micromolar concentrations of lipid A, phosphatidic acid (PA) and lysophosphatidic acid (M. G. Low and K.-S. Huang, J. Biol. Chem. 268, 8480-8490, 1993). In this study we have explored the mechanism of inhibition using synthetic analogs of lipid A, and PA. Monosaccharide analogs of lipid A, which varied in the number and position of the phosphate groups, the type of acyl group, and its linkage to the glucosamine ring, were tested for their ability to inhibit GPI-PLD. A compound (SDZ 880.431) containing 3-aza-glucosamine 1,4-diphosphate as the polar headgroup was identified which had a potency (IC(50) approximately 1 microM) similar to natural lipid A preparations. Removal of either phosphate residue increased the IC(50) markedly. Analogs of PA such as (7-nitro-2-1,3-benzoxadiazo-4-yl)amino-PA, ceramide 1-phosphate, and hexadecyl phosphate had approximately IC(50) values ranging from 1 to 5 microM, indicating that considerable variation in the structure of the hydrophobic groups was permissible. Inhibition of GPI-PLD by long-chain PA could not be blocked by high concentrations of glycerol 1-phosphate or dibutyryl PA. These results indicate that the hydrophobic groups do not have a passive role in inhibition but are directly involved in the binding interaction with GPI-PLD. We propose that this diverse group of inhibitors all bind to a common site on GPI-PLD, the central hydrophobic cavity predicted by the beta-propeller model for integrin alpha subunits and GPI-PLD.
Low
M G
MG
Department of Physiology and Cellular Biophysics, College of Physicians and Surgeons, New York, New York 10032, USA. mgL2@columbia.edu
Stütz
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D009327:4-Chloro-7-nitrobenzofurazan / Q000031:analogs & derivatives; D000818:Animals; D004305:Dose-Response Relationship, Drug; D008050:Lipid A / Q000031:analogs & derivatives*; D009005:Monosaccharides / Q000494:pharmacology*; D010712:Phosphatidic Acids / Q000494:pharmacology*; D010739:Phospholipase D / Q000037:antagonists & inhibitors*; D013329:Structure-Activity Relationship; D014346:Trypanosoma brucei brucei; D014643:Variant Surface Glycoproteins, Trypanosoma / Q000378:metabolism
D008050:Lipid A; D009005:Monosaccharides; D010712:Phosphatidic Acids; D014643:Variant Surface Glycoproteins, Trypanosoma; D010739:Phospholipase D; C065017:glycoprotein phospholipase D; D009327:4-Chloro-7-nitrobenzofurazan