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Aberrant post-translational modifications characterize the hepatic proteome of methylmalonic acidemia

异常翻译后修饰是甲基丙二酸血症肝脏蛋白质组的特征

基本信息

批准号：
10025497
负责人：
PamelaSara Elbaz Head
金额：
--
依托单位：
NATIONAL HUMAN GENOME RESEARCH INST
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-01 至 2023-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10025497
关键词：
Acyl Coenzyme A Acylation Alleles Amino Acids Antibodies Arginine Biological Assay CRISPR/Cas technology Catabolism Cell Line Cells Clinical Treatment Coenzyme A Complex Defect Development Disease Environment Enzymes Exhibits Fatty Acids Functional disorder Genetic Diseases Glutathione Hepatic Hepatocyte Histone Deacetylase Human Immunoprecipitation Impairment In Vitro Inborn Errors of Metabolism Investigation Isoleucine Knock-in Knock-out Lead Liver Liver Mitochondria Lysine Mass Spectrum Analysis Measures Metabolic Metabolic Diseases Metabolism Methionine Methylmalonic Acid Methylmalonyl-CoA Mutase Mitochondria Mitochondrial Matrix Mitochondrial Proteins Modeling Modification Morbidity - disease rate Mus Mutase Mutate Mutation Parents Pathway interactions Patients Phenotype Post-Translational Protein Processing Protein Family Proteins Proteome Protocols documentation Reaction Reagent Research Resistance Risk SIRT1 gene Sampling Severity of illness Sirtuins Site-Directed Mutagenesis Stress Surveys Symptoms Therapeutic Intervention Threonine Tissue Sample Tissues Toxic effect Tryptophan Urea Valine Vitamin B 12 Western Blotting arginyllysine disease phenotype fatty acid metabolism gene therapy genetic regulatory protein improved in vivo liver function loss of function methylmalonic aciduria methylmalonyl-coenzyme A mimetics mortality mouse model novel overexpression propionyl-coenzyme A protective effect small molecule succinyl-coenzyme A targeted treatment thioester

项目摘要

Project Summary Organic acidemias (OAs), such as methylmalonic acidemia (MMA), are a group of inborn errors of metabolism that typically arise from defects in the catabolism of amino- and fatty acids. OAs are difficult to treat and have multisystemic manifestations, leading to increased morbidity and mortality. Accretion of acyl-CoA species is postulated to cause intracellular toxicity. Here, we explore an alternative pathophysiological consequence of impaired acyl-CoA metabolism: the accumulation of aberrant posttranslational modifications (PTMs) that modify enzymes in critical intracellular pathways, especially during periods of increased stress. Using a mouse model that recapitulates the hepatic mitochondriopathy of MMA (Mmut-/-;TgINS-MCK-Mut) as well as MMA patient liver tissues, I surveyed PTMs in hepatic extracts with propionyl- and malonyl-lysine antibodies. I discovered widespread hyper-acylation in the MMA mice and MMA patient tissue samples compared to their respective control samples, but not in mice with Acsf3 deficiency, a disorder of acyl-CoA synthesis. Next, I prepared anti- PTM antibody columns, purified hepatic extracts from MMA and control mice, and performed mass spectrometry to characterize the PTM proteome. Excessive acylation of enzymes involved in glutathione, urea, arginine, lysine, tryptophan, valine, isoleucine, methionine, threonine, and fatty acid metabolism were detected in the MMA mice but not controls, and further validated mass spectrometry Cps1 hyperacylation via immunoprecipitation analysis and Western blotting. In parallel, we generated, via nonenzymatic acylation reactions, PTM-modified BSA targets for in vitro analyses. We purified, then assayed, SIRT1-7 deacylase activity using BSA-PTM standards to identify the SIRT(s) that most efficiently remove MMA related PTMs. Because PTMs usually inhibit enzyme function, our observations suggest that hyperacylation of key enzymes in pathways known to be dysregulated in MMA likely contributes to altered metabolism and identifies a new set of targets for therapeutic intervention.

项目总结