The overall goal of this project is to investigate the biochemistry, heterogeneity and molecular biology of the human cytosolic sulfotransferases (STs) and to understand the role of sulfation in the metabolism of drugs, xenobiotics and endogenous substrates in humans. The First Award involved the investigation of the phenol sulfotransferases (PST) in human platelet and liver cytosol; however, during these studies we also identified, purified and cloned the major steroid/bile acid ST in human liver, dehydroepiandrosterone (DHEA) ST. DHEA is a member of the steroid ST family of STs and is not closely related to the PSTs but also has important roles in steroid and drug metabolism. Therefore, the Specific Aims of this project are expanded to include the investigation of the protein chemistry and molecular biology of all human cytosolic STs. We believe that this fundamental biochemical information will improve our understanding of human drug metabolism and provide information for drug design and therapy, as well as increase our knowledge of the functions of sulfation in cellular physiology. Sulfation is one of the major pathways for the biotransformation and excretion of drugs and endogenous compounds, such as steroids and catecholamines. Sulfate conjugation generally results in a decrease in the biological activity of a compound and an increase in its solubility; however, sulfation may be involved in the bioactivation of compounds to mutagens and carcinogens, or to pharmacologically active forms. Most animal species appear to have multiple forms of hepatic cytosolic STs whereas, only three distinct major forms of cytosolic ST have been identified. Characterization and investigation of these enzymes will be important to understand their biological and drug metabolism functions. Identification and characterization of other forms of ST will also be necessary to enlarge our understanding of the functions of sulfation and the heterogeneity of STs. This project will focus on the two forms of phenol ST (PST) in human liver cytosol, M-PST and P-PST, and the major steroid/bile acid ST in human liver and adrenal cytosol, DHEA-ST. We will capitilize on the knowledge and important tools developed by the First Award.
The Specific Aims of this proposal are to: 1) characterize the molecular structure of the human liver PSTs and DHEA-ST; 2) investigate the localization of DHEA-ST in human tissues; 3) investigate the genomic organization and heterogeneity of allelic forms of PST in human liver; 4) investigate the ability of the PSTs and DHEA-ST to conjugate drugs and bioactive mutagens following expression in cultured cells; and 5) identify, purify , and clone human estrogen sulfotransferase.
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