Characterization of CoA-independent transacylase activity in U937 cells.
Coenzyme A-independent transacylase (CoA-IT) mediates the transfer of polyunsaturated fatty acids from the sn-2 position of a donor phospholipid to the sn-2 position of an acceptor lyso-phospholipid. We have characterized this activity in U937 cells, a human monocytic cell line. The microsomes of these cells contained CoA-IT activity which demonstrated a fatty acid preference for transferring arachidonic acid into exogenously added 1-alkyl-2-lyso-GPC. This enzymatic activity was optimum between pH 6.5 and 9, was heat labile and displayed an apparent Km for 1-alkyl-2-lyso-GPC of 0.4 microM. This activity was not dependent on Ca2+, Mg2+, CoA or ATP, was not inhibited by 2-mercaptoethanol nor by addition of product, 1-alkyl-2-acyl-GPC. The activity of this enzyme was not altered by differentiation of U937 cells towards the macrophage with Me2SO. Treatment of U937 cells with dexamethasone had no effect on transacylase activity. The activity of this enzyme was decreased by the serine esterase inhibitors phenylmethyl-sulfonyl fluoride and N-tosyl-L-phenylalanine chloromethyl ketone and by the histidine modifier diethyl pyrocarbonate, suggesting that CoA-IT may belong to a family of acyltransferase enzymes typified by LCAT. CoA-IT activity was not affected by compounds that affect PLA2 activity, such as quinacrine, aristolochic acid and arachidonic acid, suggesting a mechanism of action for CoA-IT different from classical, low molecular weight PLA2 enzymes. In conclusion, U937 cells contain CoA-IT activity and this study extends our previous knowledge of this enzyme by demonstrating the differences between CoA-IT and PLA2 enzymes and suggesting similarities between CoA-IT and LCAT.
Winkler
J D
JD
Department of Pharmacology, SmithKline Beecham Pharmaceuticals, King of Prussia, PA 19406.
Sung
C M
CM
Bennett
C F
CF
Chilton
F H
FH
D000217:Acyltransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000378:metabolism; D034341:Aristolochic Acids; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D003907:Dexamethasone / Q000494:pharmacology; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008861:Microsomes / Q000201:enzymology*; D010616:Phenanthrenes / Q000494:pharmacology; D011796:Quinacrine / Q000494:pharmacology
D001095:Arachidonic Acids; D034341:Aristolochic Acids; D010616:Phenanthrenes; D016718:Arachidonic Acid; D003907:Dexamethasone; C000228:aristolochic acid I; D004492:Edetic Acid; D000217:Acyltransferases; C039766:arachidonyl transacylase; D011796:Quinacrine; D004047:Diethyl Pyrocarbonate