Mechanisms and Functions of Human Sulfotransferases

描述（由申请人提供）：II期药物代谢酶磺基转移酶（SULTs）催化的硫酸化在调节不同激素和药物和其他外源性药物的解毒中很重要。磺化还会导致致癌物原的生物活化，从而产生毒性作用。本研究项目的长期目标是了解人体SULT的生物学功能，并研究其在生理和病理条件下与人体健康的相关性。本提案的具体目的如下：1。研究人类SULTs的催化、底物抑制和产物抑制/激活机制。本文将利用动力学分析、同位素交换、硫酸酸化活性位点氨基酸残基鉴定和位点定向诱变等方法对所提出的旁路有序机制和相关替代机制进行研究。这些信息将对生物转化途径的预测具有重要意义。2. 目的：探讨磺化药物对人体SULT催化活性的影响。将研究临床重要药物硫酸盐对人SULTs催化活性的抑制和激活作用。大肠杆菌表达和纯化人SULTs；人肠细胞质；人类Hep G2和Caco-2细胞将用于这些研究。临床药物对人体SULT活性的影响可能会干扰SULT正常的激素调节和外源解毒等生物学功能。3. 明确人SULT1E1的氧化调控机制。我们将利用酶分析、Western blot、RT-PCR、氨基酸修饰、定点诱变、动力学分析、晶体结构分析和计算机建模等方法，研究人SULT1E1在Hep G2和Caco-2细胞中的氧化调节和纯化人SULT1E1的氧化还原硫醇调节机制。了解某些人体SULT的氧化调节对于了解SULT在生理和病理条件下的功能是很重要的。本提案研究人类的结果。这些研究将对了解SULT的生物学功能具有重要意义，包括激素调节、药物代谢、外源解毒和前致癌物的生物活化。这些知识对于了解药物副作用、药物相互作用、药物开发以及SULTs在癌症预防和病因方面的潜在作用非常重要。

DESCRIPTION (provided by applicant): Phase II drug metabolizing enzymes sulfotransferases (SULTs) catalyzed sulfation is important in the regulation of different hormones and the detoxification of drugs and other xenobiotics. Sulfation also leads to bioactivation of procarcinogens leading to toxic effect. The long-term goal of this research project is to understand human SULT biological functions and to investigate their relevance to human health under physiological and pathological conditions. Specific aims in this proposal are as follows: 1. To investigate mechanisms of catalysis, substrate inhibition, and product inhibition/activation of human SULTs. The proposed bypass ordered mechanism and related alternative mechanisms will be investigated using kinetic analysis, isotope exchange, sulfated active site amino acid residue identification, and site-directed mutagenesis. The information will have important implications for the prediction of biotransformation pathways. 2. To investigate the effect of sulfated drugs on human SULT catalytic activities. The inhibition and activation effect of clinically important drug sulfates on catalytic activities of human SULTs will be investigated. E. coli expressed and purified human SULTs; human intestinal cytosols; and human Hep G2 and Caco-2 cells will be used for these investigations. The effect of clinical drugs on human SULT activities may interfere SULT normal biological functions in hormone regulation and xenobiotic detoxification. 3. To define oxidative regulation mechanisms of human SULT1E1. Oxidative regulation of human SULT1E1 in Hep G2 and Caco-2 cells and redox thiol regulation mechanisms of purified human SULT1E1 will be investigated using enzyme assay, Western blot, RT-PCR, amino acid modification, site-directed mutagenesis, kinetic analysis, crystal structure analysis, and computer modeling methods. Knowledge on oxidative regulation of certain human SULT is important in understanding the ability of SULT functioning under physiological and pathological conditions. This proposal studies human SULTs. These studies will be significant in understanding SULT biological functions including hormone regulation, drug metabolism, xenobiotic detoxification and procarcinogen bioactivation. The knowledge will be important in understanding drug side effect, drug-drug interaction, drug development, and the potential roles SULTs play in cancer prevention and causation.