ARYL AND ALCOHOL SULFOTRANSFERASES IN DRUG METABOLISM

这项研究的长期目标是更好地理解和更多 可靠地预测芳基转移酶(AST)IV和酒精的作用 磺基转移酶在细胞毒性、致突变性和/或致癌性中的作用 对药物、致癌物和其他外源物质的反应。准确的 涉及磺基转移酶的代谢反应的预测是 对于拥有或被转化的分子来说尤其重要 代谢产物中含有苯甲醇、芳基异羟肟酸和N- 羟基芳胺官能团，因为这些官能团的硫酸盐化 基团通常是化学反应形成的关键步骤 能够与细胞形成共价键的代谢物 大分子。这种与关键细胞分子的共价键是 第一步导致各种细胞毒性，免疫学， 致突变和致癌反应。这项研究中提出的 应用是基于这样一个前提，即 磺基转移酶的催化特性和机制，以及 这些酶在特定细胞内的定位和调节 肝脏等组织对于准确预测 细胞内大分子的共价改变的可能性 单个细胞。 天冬氨酸转氨酶IV的定量构效关系研究 和sta将集中在疏水和空间性质。 与这些酶结合和催化所需的。这些研究将 还提供了普遍适用的量化比较 两种磺基转移酶的立体选择性 系列手性底物。该项目的第二个具体目标是 识别和定位特定的氨基酸残基，这些氨基酸残基对 天冬氨酸转氨酶IV和天冬氨酸转氨酶的底物结合及催化活性。这 两项调查都继续成功地使用核糖核苷酸 双醛亲和标记剂用于分析土壤中的残留 3‘-磷酸-腺苷-5’-磷酸-硫酸盐的结合部位及使用新方法 硫基受体上残留物的测定方法 地点。这些新方法将利用一系列同源的底物- 含有溴乙酰胺基团的类似物作为AST IV的亲和标记。 这些亲和力标签旨在提供有关亲缘关系的信息 特定氨基酸残基离硫基位置的距离 转移到酶上。这项研究的第三个主要组成部分是 天门冬氨酸氨基转移酶IV和天冬氨酸转氨酶异质性的基础探讨 大鼠肝小叶内肝细胞的原位杂交研究 用免疫组织化学分析。这些调查将提供 关于这两种类型的关系的重要新信息 肝脏中的磺基转移酶。

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.