Copper (Cu) is a transition metal which is essential to human health; it helps maintain healthy bones, blood vessels, nerves, and immune function. Regulation of copper transport into the cell membrane is controlled by Ctr1, which is a plasma membrane protein that possesses a high affinity for copper at its N-terminus. The acquisition of copper by this protein can be disrupted by reactive oxygen species (ROS), which are free radicals that react with the protein easily and cause damage leading to neurodegenerative disorders such as Alzheimer’s, Parkinson’s, and other health conditions including osteoporosis and oxidative stress. Nuclear magnetic resonance (NMR) can be used to see the interactions between Ctr1 and ROS. NMR can also assist with understanding which residues are interacting with copper; this information is necessary to understand the function of the protein. However, chemical synthesis of a protein with 13C and 15N isotopes, which are required for NMR, is costly and produces a large amount of chemical waste. As an alternative to peptide synthesis, we expressed two forms of the N-terminal domain of Ctr1 as SUMO fusion proteins in Escherichia coli which could be isotopically labeled. This process minimized the cost and amount of hazardous waste associated with the production of these proteins for analysis.

Optimization of the expression and purification of the copper transport protein for NMR analysis

Macromolecular Chemistry: The Second Century

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ACS Spring 2020 National Meeting & Expo

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.