Substrate specificity of THCA-CoA oxidases from rat liver light mitochondrial fractions on dehydrogenation of 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestanoic acid CoA thioester

Steroids. 1998 Nov;63(11):603-7. doi: 10.1016/s0039-128x(98)00070-1.

Abstract

The substrate specificity of rat liver peroxisomal 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestanoyl-CoA (THCA-CoA) oxidases, which catalyze the dehydrogenation of 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestanoic acid (THCA) CoA thioester, having an asymmetric center at C-25, to form (24E)-3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholest-24-enoic acid (delta 24-THCA) CoA thioester, was studied. The stable isotope labeled substrates, [3,7,12-18O3]-(25R)- and (25S)-THCA CoA thioesters were synthesized by an exchange reaction of carbonyl oxygens on a steroid nucleus of 3,7,12-trioxo-5 beta-cholestanoic acid, followed by metal hydride reduction and condensation reaction with CoA. After incubation of a mixture of unlabeled (25R)- and 18O-labeled (25S)-THCA CoA thioester, or vice versa, with hepatic peroxisomal THCA-CoA oxidases, biotransformed delta 24-THCA was determined by liquid chromatography/atmospheric pressure chemical ionization mass spectrometry. The delta 24-THCA was derived only from (25S)-THCA CoA thioester, indicating that the 25S epimer of THCA is a preferential substrate on dehydrogenation by THCA-CoA oxidases.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cholestanols / metabolism*
  • Male
  • Mass Spectrometry
  • Mitochondria, Liver / enzymology*
  • Oxidoreductases / metabolism*
  • Rats
  • Rats, Wistar
  • Substrate Specificity

Substances

  • 3alpha,7alpha,12alpha-trihydroxy--5beta-cholestanoic acid coenzyme A thioester
  • Cholestanols
  • Oxidoreductases
  • trihydroxycholestanoyl-CoA oxidase