2-Acylglycerolphosphoethanolamine acyltransferase/acyl-acyl carrier protein synthetase is a membrane-associated acyl carrier protein binding protein.
Two enzymatic activities, 2-acylglycerolphosphoethanolamine (2-acyl-GPE) acyltransferase and acyl-acyl carrier protein (acyl-ACP) synthetase, were solubilized and purified from Escherichia coli membranes. Electrophoretic analysis of the final product of the purification procedure revealed a single protein species with an apparent molecular mass of 27 kilodaltons. The ratio of acyltransferase to synthetase activities remained the same throughout the purification scheme indicating that both activities were catalyzed by the same enzyme. 2-Acyl-GPE acyltransferase exhibited an apparent ACP Km of 64 nM under standard assay conditions that increased to 10 microM when the assay was conducted in the presence of 0.4 M LiCl. Acyl-ACP synthetase activity was not detected in the absence of 0.4 M LiCl, and the apparent ACP Km for this reaction was 16 microM. Direct evidence that ACP was a subunit of the acyltransferase/synthetase was obtained by the adsorption of both catalytic activities to an ACP-Sepharose affinity column and by the binding of [3H]ACP to the purified enzyme preparation. The apparent Km for acyl-ACP was 13 microM, and the rate of acyl transfer from this acyl donor was enhanced by the addition of 0.4 M LiCl indicating that the exchange of enzyme-bound ACP for acyl-ACP was a determinant factor in the rate of phosphatidylethanolamine formation from acyl-ACP. These data indicate that the 2-acyl-GPE acyltransferase and acyl-ACP synthetase reactions are catalyzed by the same membrane protein that possesses a high affinity binding site for soluble ACP.
Cooper
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CL
Department of Biochemistry, St. Jude Children's Research Hospital, Memphis, Tennessee 38101.
Hsu
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Jackowski
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Rock
C O
CO
D000213:Acyl Carrier Protein / Q000378:metabolism*; D000217:Acyltransferases / Q000302:isolation & purification / Q000378:metabolism*; D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism*; D002712:Chlorides / Q000494:pharmacology; D002846:Chromatography, Affinity; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D008094:Lithium / Q000494:pharmacology; D018021:Lithium Chloride; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008970:Molecular Weight; D010714:Phosphatidylethanolamines / Q000378:metabolism; D010770:Phosphotransferases / Q000302:isolation & purification / Q000378:metabolism*; D012995:Solubility; D017855:Transferases (Other Substituted Phosphate Groups)
D000213:Acyl Carrier Protein; D001426:Bacterial Proteins; D002712:Chlorides; D008565:Membrane Proteins; D010714:Phosphatidylethanolamines; D008094:Lithium; D000217:Acyltransferases; C041419:acyl-(acyl-carrier-protein)-phospholipid acyltransferase; D010770:Phosphotransferases; D017855:Transferases (Other Substituted Phosphate Groups); C021364:holo-(acyl-carrier-protein) synthase; D018021:Lithium Chloride