Abstract
An in vitro study of bile acid-CoA:amino acid N-acyltransferase activity of rat liver was undertaken in order to determine whether separate amino acid-specific enzymes catalyzed the formation of glycine and taurine conjugates of bile acids as postulated by others. Polyacrylamide gel electrophoresis of 200-fold purified enzyme localized the glycine- and taurine-dependent activities to a single band. Both activities were optimal at pH 7.8 and showed similar loss of activity at pH 6.0, pH 9.0, in the presence of 5,5'-dithiobis(2-nitrobenzoic acid), and at temperatures exceeding 50 degrees. With the purified fraction, Km for glycine was 31 mM and Km for taurine was 0.8 mM. Km for several bile acid-CoA substrates was approximately 20 micron and independent of the amino acid acceptor. Only amino acids with terminal alpha- or beta-amino groups were active as acyl acceptors. Acyl donors were limited to bile acid-CoA derivatives. The data support the conclusion that the rat has a single bile acid-CoA:amino acid N-acyltransferase. The substrate kinetics are consistent with previous observations that taurine conjugates predominate in rat bile at normal hepatocellular concentrations of glycine and taurine.
Highlights
An in vitro study of bile acid-CoA:amino acid X-acyltransferase activity of rat liver was undertaken in order to determine whether separate amino acid-specific enzymes catalyzed the formation of glycine and taurine conjugates of bile acids as postulated by others
The substrate kinetics are consistent with previous observations that taurine conjugates predominate in rat bile at normal hepatocellular concentrations of glycine and taurine
Assays were run by the DCPIP reduction method with 100 mM glycine or 20 rnM taurine as amino acid acceptors
Summary
Purification and Characterization of Rile Acid-CoA: Amino Acid N-Acyltransferase from Rat Liver*. An in vitro study of bile acid-CoA:amino acid X-acyltransferase activity of rat liver was undertaken in order to determine whether separate amino acid-specific enzymes catalyzed the formation of glycine and taurine conjugates of bile acids as postulated by others. Studies of the factors which control the prevalence of the amino acid moiety of the bile acid conjugates in a given species have suggested that both amino acid availability (intrahepatic taurine synthesis) and enzyme specificity are potentially important [7, 8]. Have developed sensitive and specific methods for the determination of bile acid-CoA:amino acid N-acyltransferase activity [12] With these methods, we have been able to partially purify and study the substrate specificity of this enzyme in rat liver. The data presented in this paper support the opposite conclusion: that rat has a single bile acid-CoA:amino acid N-acyltransferase which catalyzes the formation of both taurine and glycine conjugates. The result is that at physiologic levels and glycine, taurine is the preferred substrate
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