A Recipe for Protein Footprinting

Michael Gross, professor of chemistry in Arts & Sciences and of immunology and internal medicine at the School of Medicine, and his team are experts in footprinting proteins — that is, using advanced methods for investigating the structure and interactions of proteins within larger molecules.

By sharing their method for fast photochemical oxidation of proteins (FPOP), a means of protein footprinting, they hope to support other labs in developing broader applications of FPOP to better address outstanding questions in structural biology.

“FPOP has drawn significant attention because it complements existing footprinting,” said Roger Liu, a graduate student working with Gross and first author of a new publication about protein footprinting in the journal Nature Protocols. “Its major advantages include fast labeling time frame, irreversible nature, high sensitivity and relatively broad amino acid residue coverage.”

Despite the compelling advantages of FPOP, the technical difficulty of establishing the platform has caused a lag in broader applications. Liu cites challenges including choosing the proper laser, setting up the laser optics, establishing the flow system, acquiring the footprint and analyzing the results by mass spectrometry.

“We always thought that the best way to disseminate FPOP is by applications,” Gross said. “Following its discovery, we have implemented it for problems in biochemistry and biophysics.”

Fast protein folding could be beneficial for mapping epitopes, which are the parts of an antigen that are recognized by the immune system. This application is potentially important for scientists and medical professionals racing to address the COVID-19 pandemic.

Other applications could include understanding aggregating proteins, with implications in Alzheimer’s disease; uncovering hidden conformational changes invisible to other structural methods; and determining binding sites and binding affinities of small molecules that bind to proteins.

Gross added: “Currently, in a collaboration with Weikai Li in the Department of Biochemistry and Molecular Biophysics, we are moving into transmembrane and membrane-associated proteins where new structural methods are desperately needed for this important class of proteins.”

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