Bile acids aid in the absorption of lipids by the intestine and aid in the biliary elimination of compounds that cannot be excreted in the urine. In normal bile, bile acids are present as glycine (gly) or taurine (tau) conjugates which arise as the result of hepatic metabolism. It is known that conjugation of bile acids is critical for their role in intestinal absorption and that conjugation enhances their rate of biliary excretion. We will determine if the enhanced excretion of bile acids results from an increased rate of secretion from the hepatocyte into the bile canaliculus or merely a decreased absorption from bile back into the hepatocyte. Since the solubility of cholesterol in bile is determined to some extent by the relative rates of conjugation, we want to be able to predict the relative rates of conjugation of different bile acids. To do this we will characterize the kinetic properties of the two enzymes of bile acid conjugation; the microsomal Bile Acid:CoA Ligase and the soluble Bile Acid-CoA:gly/tau N-acyltransferase. For the ligase, the reaction mechanism will have to be determined as will all of the kinetic constants for a variety of bile acids. The role of the membrane in regulating the properties of the ligase will also be examined. The N-acyltransferase will be studied by titration calorimetry to gain an understanding of its substrate specificity. The structure of a covalent substrate intermediate will also be determined in order to understand the chemistry of the active site. We are also interested in the evolutionary changes in the N-acyl-transferase. Non-mammals have large enzyme forms (65,000 daltons) that can synthesize only tau conjugates. Mammals have smaller enzymes forms (50,000 daltons) that can synthesize both gly and tau conjugates. We will determine if the mammalian form evolved from the same ancestral gene and if it did, what portion of the genome was eliminated during evolution and what precisely accounts for the evolutionary change in substrate specificity. Bile acids also are known to inhibit the detoxification of electrophilic toxins catalyzed by glutathione S-transferases. We will determine if the inhibition of these enzymes in the colon is responsible for bile acids being promoters of colon carcinogenesis. We will also determine if inhibition of these enzymes in the liver enhances the toxicity of certain hepatotoxins. Finally, we will begin studies on the glucuronidation of bile acids since this pathway is important in cholestasis.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK019212-11
Application #
3226292
Study Section
General Medicine A Subcommittee 2 (GMA)
Project Start
1979-06-01
Project End
1990-05-31
Budget Start
1986-06-01
Budget End
1987-05-31
Support Year
11
Fiscal Year
1986
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Type
Schools of Medicine
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Vessey, D A; Lee, K H; Lau, E (1997) Effect of bile acids on the growth and differentiation of cultured human keratinocytes. Skin Pharmacol 10:265-74
Vessey, D A; Kelley, M (1995) Inhibition of bile acid conjugation by cyclosporin A. Biochim Biophys Acta 1272:49-52
Kelley, M; Vessey, D A (1994) Determination of the mechanism of reaction for bile acid: CoA ligase. Biochem J 304 ( Pt 3):945-9
Kelley, M; Vessey, D A (1994) Dual role of divalent cations in the bile acid:CoA ligase catalyzed reaction. Biochim Biophys Acta 1209:51-5
Vessey, D A; Benfatto, A M; Zerweck, E et al. (1990) Purification and characterization of the enzymes of bile acid conjugation from fish liver. Comp Biochem Physiol B 95:647-52
Vessey, D A; Boyer, T D (1988) Characterization of the activation of rat liver glutathione S-transferases by nonsubstrate ligands. Toxicol Appl Pharmacol 93:275-80
Boyer, T D; Vessey, D A; Kempner, E (1986) Radiation inactivation of microsomal glutathione S-transferase. J Biol Chem 261:16963-8