Lecithin-cholesterol acyltransferase (LCAT) catalyzes transacylation of intact cholesteryl esters. Evidence for the partial reversal of the forward LCAT reaction
- Wake Forest Univ., Winston-Salem, NC (USA)
Lecithin-cholesterol acyltransferase (LCAT) catalyzes the intravascular synthesis of lipoprotein cholesteryl esters by converting cholesterol and lecithin to cholesteryl ester and lysolecithin. LCAT is unique in that it catalyzes sequential reactions within a single polypeptide sequence. In this report we find that LCAT mediates a partial reverse reaction, the transacylation of lipoprotein cholesteryl oleate, in whole plasma and in a purified, reconstituted system. As a result of the reverse transacylation reaction, a linear accumulation of (3H)cholesterol occurred during incubations of plasma containing high density lipoprotein labeled with (3H)cholesteryl oleate. When high density lipoprotein labeled with cholesteryl (14C)oleate was also included in the incubation the labeled fatty acyl moiety remained in the cholesteryl (14C)oleate pool showing that the formation of labeled cholesterol did not result from hydrolysis of the doubly labeled cholesteryl esters. The rate of release of (3H)cholesterol was only about 10% of the forward rate of esterification of cholesterol using partially purified human LCAT and was approximately 7% in whole monkey plasma. Therefore, net production of cholesterol via the reverse LCAT reaction would not occur. (3H)Cholesterol production from (3H)cholesteryl oleate was almost completely inhibited by a final concentration of 1.4 mM 5,5'-dithiobis(nitrobenzoic acid) during incubation with either purified LCAT or whole plasma. Addition of excess lysolecithin to the incubation system did not result in the formation of (14C)oleate-labeled lecithin, showing that the reverse reaction found here for LCAT was limited to the last step of the reaction. To explain these results we hypothesize that LCAT forms a (14C)oleate enzyme thioester intermediate after its attack on the cholesteryl oleate molecule.
- OSTI ID:
- 7027320
- Journal Information:
- Journal of Biological Chemistry; (USA), Journal Name: Journal of Biological Chemistry; (USA) Vol. 265:5; ISSN JBCHA; ISSN 0021-9258
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
ACYLATION
CARBON 14 COMPOUNDS
CARBOXYLIC ACIDS
CHEMICAL REACTIONS
CHOLESTEROL
DOUBLE LABELLING
ENZYME ACTIVITY
ENZYMES
ESTERIFICATION
ESTERS
HYDROGEN COMPOUNDS
HYDROXY COMPOUNDS
ISOTOPE APPLICATIONS
ISOTOPE DILUTION
LABELLED COMPOUNDS
LABELLING
METABOLISM
MONOCARBOXYLIC ACIDS
OLEIC ACID
ORGANIC ACIDS
ORGANIC COMPOUNDS
STEROIDS
STEROLS
TRACER TECHNIQUES
TRANSFERASES
TRITIUM COMPOUNDS