Chemical synthesis of bile acid acyl-adenylates and formation by a rat liver microsomal fraction |
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Authors: | Shigeo Ikegawa Hiromi Ito Masako Maeda Kuniko Mitamura |
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Institution: | a Faculty of Pharmaceutical Sciences, Kinki University, 3-4-1 Kowakae, Higashi-osaka, 577-8502, Japan b School of Pharmaceutical Sciences, Showa University, 1-5-8, Hatanodai, Shinagawa, Tokyo, 142-8555, Japan c Department of Medicine, University of California, San Diego, La Jolla, CA 92093-0063, United States |
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Abstract: | In mammals, unconjugated bile acids formed in the intestine by bacterial deconjugation are reconjugated (N-acylamidated) with taurine or glycine during hepatocyte transport. Activation of the carboxyl group of bile acids to form acyl-adenylates is a likely key intermediate step in bile acid N-acylamidation. To gain more insight into the process of bile acid adenylate formation, we first synthesized the adenylates of five common, natural bile acids (cholic, deoxycholic, chenodeoxycholic, ursodeoxycholic, and lithocholic acid), and confirmed their structure by proton NMR. We then investigated adenylate formation by subcellular fractions of rat liver (microsomes, mitochondria, cytosol) using a newly developed LC method for quantifying adenylate formation. The highest activity was observed in the microsomal fraction. The reaction required Mg2+ and its optimum pH was about pH 7.0. In term of maximum velocity (Vmax) and the Michaelis constant (Km), the catalytic efficiency of the enzyme under the conditions used was highest with cholic acid of the bile acids tested. The formation of cholyl-adenylate was strongly inhibited by lithocholic and deoxycholic acid, as well as by palmitic acid; ibuprofen and valproic acid were weak inhibitors. In cholestatic disease, such adenylate formation might lead to subsequent bile acid conjugation with glutathione or proteins. |
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Keywords: | Cholyl-adenylate (CA-AMP) cholyl adenosine 5&prime -phosphate diester Deoxycholyl-adenylate (DCA-AMP) deoxycholyl adenosine 5&prime -phosphate diester Ursodeoxycholyl-adenylate (UDCA-AMP) ursodeoxycholyl adenosine 5&prime -phosphate diester Lithocholyl-adenylate (LCA-AMP) lithocholyl adenosine 5&prime -phosphate diester 12-Oxolithocholyl-adenylate (12-oxoLCA-AMP) 12-oxolithocholyl adenosine 5&prime -phosphate diester Valproyl-adenylate (valproyl-AMP) valproyl adenosine 5&prime -phosphate diester Ibuprofenyl-adenylate (ibuprofenyl-AMP) ibuprofenyl adenosine 5&prime -phosphate diester AMP adenosine 5&prime -monophosphate ATP adenosine-5&prime -triphosphate Palmitic acid hexadecanoic acid Valproic acid 2-propylpentanoic acid Ibuprofen 2-[4-(2-methylpropyl)phenyl]propanoic acid |
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