Substrate specificities of 3-oxoacyl-CoA thiolase A and sterol carrier protein 2/3-oxoacyl-CoA thiolase purified from normal rat liver peroxisomes - Sterol carrier protein 2/3-oxoacyl-CoA thiolase is involved in the metabolism of 2-methyl-branched fatty acids and bile acid intermediates

摘要: The two main thiolase activities present in isolated peroxisomes from normal rat liver were purified to near homogeneity. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the first enzyme preparation displayed a single band of 41 kDa that was identified as 3-oxoacyl-CoA A (thiolase A) by N-terminal amino acid sequencing, second consisted 58- and 46-kDa band, 58-kDa polypeptide reacted with antibodies raised against either sterol carrier protein 2 or domain 2/3-oxoacyl-CoA (SCP-2/thiolase), formerly also called X, whereas only domain, Internal peptide sequencing confirmed is SCP-2/thiolase SCP-2/thiolase. Thiolase catalyzed cleavage short, medium, long straight chain 3-oxoacyl-CoAs, medium 3-oxoacyl-CoAs being best substrates, inactive 2-methyl-branched 3-oxo-2-methylpalmitoyl-CoA bile intermediate 24-oxo-trihydroxycoprostanoyl-CoA. active but intermediate. In peroxisomal extracts, more than 90% activity toward associated A. Kinetic parameters (K-m V-max) determined for each different substrates. Our results indicate following: 1) (main) thiolases are SCP-2/thiolase; 2) responsible thiolytic 3-oxoacyl-CoAs; 3) derivatives fatty acids side cholesterol.