Acylglycerol reactivity and reaction mechanism of bovine milk lipoprotein lipase.
The sequential lipolysis of trioleoylglycerol and the triacylglycerols of very-low-density lipoprotein by bovine milk lipoprotein lipase can be described by the consecutive reactions: (formula: see text) where k'1, k'2 and k'3 are apparent first-order rate constants. The values of these rate constants dictate several conclusions concerning the reaction mechanism of this enzyme. The significant differences in the k'1, k'2 and k'3 values for trioleoylglycerol substrate imply that cleavage of the acyl-enzyme intermediate is not the rate-limiting step of the overall lipolysis reaction. This conclusion is further supported by the lack of an effect of hydroxylamine on the reaction rate. In addition, the observed isotope effect of k1 (H2O): k1(D2O) of 1.32 with trioleoylglycerol substrate suggests that the acyl-enzyme formation may contribute to the rate-limiting step of the lipoprotein-lipase-catalyzed reaction. In the presence of excess bovine serum albumin, the transfer of fatty acid product from the enzyme to albumin must be fast, since the k'1 values are not dependent on albumin concentration. When albumin is not in excess, the reaction is retarded and the study of reaction kinetics demonstrates negligible reaction after the available albumin is saturated.
Wang
C S
CS
Lipoprotein and Atherosclerosis Research Program, Oklahoma Medical Research Foundation, Oklahoma City 73104.
Hartsuck
J A
JA
Downs
D
D
Bass
H A
HA
D000818:Animals; D002417:Cattle; D004075:Diglycerides / Q000378:metabolism; D005989:Glycerides / Q000378:metabolism*; D006868:Hydrolysis; D006898:Hydroxylamines / Q000494:pharmacology; D007700:Kinetics; D008071:Lipoprotein Lipase / Q000378:metabolism*; D008079:Lipoproteins, VLDL / Q000097:blood / Q000378:metabolism; D008892:Milk / Q000201:enzymology*; D012710:Serum Albumin, Bovine / Q000494:pharmacology; D014280:Triglycerides / Q000378:metabolism
D004075:Diglycerides; D005989:Glycerides; D006898:Hydroxylamines; D008079:Lipoproteins, VLDL; D014280:Triglycerides; D012710:Serum Albumin, Bovine; D008071:Lipoprotein Lipase