ARYL AND ALCOHOL SULFOTRANSFERASES IN DRUG METABOLISM

点击翻译按钮获取中文摘要

The long-term objective of this research is to better understand and more reliably predict the roles that aryl sulfotransferase (AST) IV and alcohol sulfotransferase STa play in the cytotoxic, mutagenic and/or carcinogenic responses to drugs, carcinogens, and other xenobiotics. The accurate prediction of metabolic reactions involving sulfotransferases is particularly important for molecules that either possess or are converted into metabolites containing benzylic alcohol, arylhydroxamic acid, and N- hydroxy arylamine functional groups, since sulfation of these functional groups is often the critical step in the formation of chemically reactive metabolites that are capable of forming covalent bonds with cellular macromolecules. Such Covalent bonding with critical cellular molecules is the initial step leading to a variety of cytotoxic, immunologic, mutagenic, and carcinogenic responses. The research proposed in this application is based on the premise that quantitative analyses of the catalytic specificities and mechanisms of sulfotransferases, as well as the localizations and regulation of these enzymes within specific cells in tissues such as the liver, are essential for accurately predicting the potential for covalent alteration of cellular macromolecules within individual cells. Investigations on quantitative structure-activity relationships for AST IV and STa will be focused on the hydrophobic and steric characteristics required for binding and catalysis with these enzymes. These studies will also provide generally applicable quantitative comparisons of the stereoselectivity of the two sulfotransferases through use of a homologous series of chiral substrates. A second specific aim of this project is to identify and locate specific amino acid residues that contribute to the substrate binding and catalytic activity of AST IV and STa. This investigation both continues the successful use of ribonucleotide dialdehyde affinity labeling reagents for elucidation of residues at the binding site for 3'-phosphoadenosine 5'-phosphosulfate and employs new approaches for the determination of residues at the sulfuryl acceptor site. These new methods will utilize a homologous series of substrate- analogs containing bromoacetamido groups as affinity labels for AST IV. These affinity labels are designed to provide information on the relative distances of specific amino acid residues from the site of sulfuryl transfer on the enzyme. The third major component of this research is an exploration of the basis for heterogeneity of AST IV and STa within hepatocytes across the rat liver lobule by combining in situ hybridization with immunohistochemical analyses. These investigations will provide important new information on the relation of these two types of sulfotransferases within liver.