Enzymology of Post-translational Modifications

翻译后修饰的酶学

基本信息

批准号：
7888852
负责人：
CAROL A FIERKE
金额：
$ 37.33万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
财政年份：
1988
资助国家：
美国
起止时间：
1988-07-01 至 2014-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7888852
关键词：
Acetylation Active Sites Address Affect Affinity Affinity Chromatography Amino Acids Antifungal Agents Benzophenones Binding Biological Biological Process C-terminal Candida albicans Cardiovascular Diseases Catalysis Cell Differentiation process Cell physiology Cells Chemicals Chimeric Proteins Clinical Clinical Trials Code Complement Cysteine Cysteine-Rich Domain Deacetylase Deacetylation Dependence Development Diabetes Mellitus Dimethylallyltranstransferase Disease Dissociation Drug Design Enzymatic Biochemistry Enzymes Gel Gene Expression Goals Histone Acetylation Histone Deacetylase Histone Deacetylase Inhibitor Histones Homologous Gene Hydrolysis In Vitro Individual Isoenzymes Kinetics Label Lysine Malignant Neoplasms Mass Spectrum Analysis Measurement Membrane Metals Modification Molecular Mutagenesis N-acetylglucosamine deacetylase Oxidation-Reduction Palmitoyl Coenzyme A Pathway interactions Peptides Phenylalanine Post-Translational Protein Processing Protein Acetylation Protein Isoprenylation Proteins Proteolysis RNA Interference Reaction Regulation Regulatory Pathway Relative (related person)Role Site Site-Directed Mutagenesis Specificity Stress Structure Substrate Specificity Transferase Yeasts base carboxymethylation cofactor crosslink drug development enzyme activity enzyme substrate fluorophore histone acetyltransferase in vivo inhibitor/antagonist isoprenoid nervous system disorder new therapeutic target novel palmitoylation pathogen prenylation protein farnesyltransferase protein function protein geranylgeranyltransferase protein transport public health relevance response small molecule thioester

项目摘要

DESCRIPTION (provided by applicant): Post-translational modifications are essential regulators of protein function in cells. In many cases, the mechanisms and substrate specificities of enzymes that catalyze these modifications are not well understood, despite their involvement in a multitude of diseases, including cancer, diabetes, cardiovascular disease and neurological disorders. Our studies will focus on two classes of post-translational modifications, lipidation (including prenylation and palmitoylation) and acetylation, and the enzymes that regulate these modifications. The majority of protein prenylation, whereby an isoprenoid group is appended near the C-terminus of a target protein, is catalyzed by protein farnesyltransferase (FTase) and protein geranylgeranyltransferase type I (GGTase-I). Palmitoylation is catalyzed by protein palmitoyl transferases (PATs), such as Akr1p, and is a thioester exchange reaction between palmitoyl-CoA and a cysteine on a substrate protein to form a palmitoyl thioester. Both prenylation and palmitoylation target proteins to membranes and are essential for protein trafficking. Protein acetylation is regulated by competition between histone acetyltransferases, that catalyze protein N-acetylation, and histone deacetylases (HDACs), that catalyze hydrolysis of 5-N-acetyl lysine. Acetylation affects multiple cellular processes, including gene expression and cell differentiation. All of these enzymes are current targets for drug development for a variety of diseases; HDAC inhibitors are approved for clinical use and FTase and GGTase inhibitors are currently in clinical trials. However, the full complement of in vivo substrates for these enzymes is unknown, and the functional basis for substrate selectivity by each enzyme is undefined. Moreover, the effects of these modifications on protein function and cellular pathways are frequently unclear. Therefore, significant questions remain for each enzyme, including their in vivo specificity and regulation. Here, we propose to: (1) investigate determinants of prenyltransferase substrate specificity for mammalian and Candida albicans enzymes and analyze the effects of individual prenylation pathway modifications on protein localization and function; (2) investigate the catalytic mechanism of the protein palmitoyltransferase Akr1p; (3) investigate the metal selectivity of deacetylases HDAC8 and LpxC in vivo and in vitro and the potential for regulation by metal switching in response to metal availability and redox stress; and (4) identify substrates and evaluate determinants of HDAC8 and HDAC11 substrate specificity, both in vitro and in vivo, by chemical crosslinking and activity-based probes. Our long-term goals are: (1) to determine the catalytic mechanism and molecular basis of specificity for each of these post-translational modifying enzymes; and (2) to examine the extent, biological function and regulation of prenylation, palmitoylation, and acetylation in the cell. Overall, our studies will assist in targeted drug design by describing the specific interactions that dictate substrate specificity and by identifying novel regulatory pathways as targets for disease therapies. PUBLIC HEALTH RELEVANCE: Post-translational modifications are essential regulators of protein function. Protein lipidation and acetylation are implicated in a multitude of diseases, including cancer, cardiovascular disease and neurological disorders. Identification of the protein substrates and downstream biological function will enhance the development of new therapeutics targeting these pathways.