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Revolutionizing Protein Higher Order Structural Analysis with Flash Oxidation (Fox) Protein Footprinting

Description

This chalk talk will offer an overview of Hydroxyl Radical Protein Footprinting, focusing on explaining the fundamentals of the technique and applications relevant to the pharmaceutical sciences. The foundations of hydroxyl radical protein labeling chemistry and the relationship of labeling to protein topography will be reviewed. Furthermore, a brief description of the Flash Oxidation (Fox) technology and the method used to generate radicals as well as the method for real-time dosimetry and the requirements for sample analysis by LC-MS/MS will be presented. Dr. Sharp will present examples of applications, focusing on the analysis of biopharmaceuticals, along with presenting data highlighting higher order structure effects of formulations. Dr. Sharp will show the effects of different buffers on the aggregation propensity and aggregation interfaces of a therapeutic monoclonal antibody along with verifying the ability of polysorbate to prevent aggregation without changing the topography of the monomeric protein. He will also present results defining the epitope of a therapeutic antibody. These applications illustrate a few of the potential applications of Fox technology towards the analysis of protein biopharmaceuticals.


Contributors

  • Joshua Sharp, Ph.D.

    Dr. Joshua Sharp is a world renowned and internationally recognized expert in Hydroxyl Radical Protein Footprinting (HRPF). He has over twenty-one peer-reviewed articles and has presented numerous invited lectures on the subject of biopharmaceutical HRPF.  Along with acting as the CTO of GenNext, Dr. Sharp is also an Assistant Professor of Pharmacology in the Department of BioMolecular Sciences. Dr. Sharp received his PhD from a joint program between Oak Ridge National Laboratory and the University of Tennessee under the direction of Dr. Robert Hettich in 2003. Dr. Sharp received his postdoctoral training at the National Institute of Environmental Health Sciences in the laboratory of Dr. Kenneth Tomer. Following his postdoctoral experiences, Dr. Sharp took a research faculty position in the Complex Carbohydrate Research Center in 2007. In 2015 he joined the Department of BioMolecular Sciences as an Assistant Professor.

    Dr. Sharp is active in the development and application of new mass spectrometry-based technologies for studying the structure-function relationships of proteins and carbohydrates of biomedical interest. Current applications focus on the study of interactions between neutralizing antibodies and the glycoprotein coat of HIV; characterization of chemokines and the structural factors mediating their oligomerization and function; and the identification of glycosaminoglycan structures with potential biomedical applications for a wide variety of fields including anti-microbial agents, anti-cancer therapies, and anti-inflammatory therapies.

    Dr. Sharp has published widely in the field of biomolecular mass spectrometry, pioneering some of the early work in the development of hydroxyl radical protein footprinting for studying protein structure and interactions. He has received funding from the National Institutes of Health to support his work in developing and applying new methods to study the structure of proteins, carbohydrates, and the complexes they form.

June 17, 2021
Thu 1:15 PM EDT

Duration 0H 30M

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