Examination of the role of thiolimidate formation in the cleavage of acetoacetyl-CoA catalyzed by thiolase I from porcine heart.
The potential contribution of thiolimidate formation to the increased kinetic acidity of the alpha-proton of acetyl-CoA in the carbon-carbon bond forming reaction catalyzed by 3-ketoacyl-CoA thiolase (thiolase I) from porcine heart was assessed by chemical modification and isotope exchange experiments. Thiolase is only partially inactivated after the chemical modification of lysine residues by reductive methylation, pyridoxal phosphate, or o-phthaldehyde (specific for vicinal lysine and cysteine). The thiolase-catalyzed formation of acetyl-CoA from acetoacetyl-CoA and CoASH in 18OH2 is not accompanied by the appearance of 18O in the acetyl-CoA product. These experiments effectively rule out participation of thiolimidate formation in the thiolase reaction. Other mechanisms must be employed to facilitate the abstraction of the alpha-proton of acetyl-CoA by thiolase I.
Izbicka
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Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, Houston, Texas 77030.
Gilbert
H F
HF
D000105:Acetyl Coenzyme A / Q000031:analogs & derivatives* / Q000378:metabolism; D000101:Acetyl-CoA C-Acetyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000123:Acetyltransferases / Q000378:metabolism*; D000214:Acyl Coenzyme A; D000818:Animals; D001665:Binding Sites; D006868:Hydrolysis; D008239:Lysine; D009206:Myocardium / Q000201:enzymology*; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship; D013552:Swine
D000214:Acyl Coenzyme A; C010667:acetoacetyl CoA; D000105:Acetyl Coenzyme A; D000123:Acetyltransferases; D000101:Acetyl-CoA C-Acetyltransferase; D008239:Lysine