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Elucidating the Molecular Basis of Cellular Metal Stress by using Mass Spectrometry-Based Proteomic Methods

使用基于质谱的蛋白质组学方法阐明细胞金属应力的分子基础

基本信息

批准号：
10600028
负责人：
Michael C Fitzgerald
金额：
$ 45万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-05 至 2026-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10600028
关键词：
Address Affect Binding Biological Biological Assay Biophysics Cancer cell line Candida albicans Cell Line Cell Separation Cell Survival Cell physiology Cells Chemicals Collaborations Copper Data Development Environment Escherichia coli Escherichia coli Proteins Event Exposure to Foundations Functional disorder Global Change Goals Growth Homeostasis Human Immune system In Vitro Ionophores Ions Laboratories Link Location Malignant Neoplasms Mass Spectrum Analysis Measurement Measures Metabolism Metal exposure Metalloproteins Metals Methodological Studies Methods Molecular Monitor Neurodegenerative Disorders Nutrient Nutritional Requirements Organism Outcome Outcome Study Pathway interactions Physiologic pulse Precipitation Predisposition Proteins Proteolysis Proteome Proteomics Proxy Publishing Research Signal Transduction Stress Techniques Testing Therapeutic Time Toxic effect Trace Elements Translations Work anti-cancer antimicrobial biophysical properties cancer cell cell type cofactor design experience microbial pathogen pathogenic microbe pharmacologic prostate cancer cell protein folding protein function protein misfolding tool trafficking

项目摘要

PROJECT SUMMARY/ABSTRACT Metal ions are required nutrients for cellular function, but can also be toxic if misregulated. Our immune system leverages this dichotomy by deploying mechanisms both to withhold nutrient metals from pathogens as well as overwhelm them with toxic levels, particularly of Cu and Zn. Metal imbalances at the cellular level have also been implicated in neurodegenerative diseases, and are being investigated as possible anticancer strategies. But what exactly are the targets and mechanisms of cellular metal stress? The research proposed here explores this question of cellular metal stress by seeking to identify protein targets of aberrant metal interactions by measuring global changes in protein stability across the proteome. The proposed work builds on preliminary and recently published results from this collaborative team showing the utility of a pulse proteolysis mass spectrometry method developed in co-PI Fitzgerald’s laboratory to identify protein targets of Cu in E. coli, establishing these proteomic methodologies for the study of metal-protein interactions. The overall objective of the current application is to identify proteins that are functionally affected when cells experience stress induced by exposure to excess levels of Zn and Cu. This objective will be met by using a powerful combination of mass spectrometry-based proteomic methods to address four specific aims: 1) Determine global profiles of protein stability changes as a function of cellular metal overload and metal deficiency across bacterial, fungal, and human cancer cells; 2) Establish a mechanistic basis linking differential stability of Aim 1 proteins to function; 3) Identify the relative sensitivity of proteins across the proteome to misfolding induced by Cu and Zn binding; and 4) Establish a biophysical basis for understanding the relative sensitivity of proteins to metal-induced misfolding. Understanding how protein stability is impacted across the proteome upon exposure to normal or aberrant levels of Cu and Zn has important implications for understanding metal-induced toxicity and mechanisms cells use to maintain metal homeostasis in the face of metal-associated stress. By studying proteomes from microbial, fungal, and human cancer cells, the outcomes of these studies will advance our understanding of how these organisms respond at the proteome level to changing metal environments imposed by the host immune system. These studies will inform and impact the development of pharmacological agents against microbial pathogens and cancer cells, and align with the applicant’s long-term goals to develop chemical tools to manipulate biological metal ion location, speciation, and reactivity for potential therapeutic benefit.

项目总结/文摘