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

> **NIH NIH R01** · DUKE UNIVERSITY · 2022 · $439,733

## Abstract

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.

## Key facts

- **NIH application ID:** 10467488
- **Project number:** 1R01GM145035-01A1
- **Recipient organization:** DUKE UNIVERSITY
- **Principal Investigator:** Michael C Fitzgerald
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $439,733
- **Award type:** 1
- **Project period:** 2022-04-05 → 2026-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10467488

## Citation

> US National Institutes of Health, RePORTER application 10467488, Elucidating the Molecular Basis of Cellular Metal Stress by using Mass Spectrometry-Based Proteomic Methods (1R01GM145035-01A1). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10467488. Licensed CC0.

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