Modification of Proteins and Peptides with Free Radical Precursors

University Of Southern Mississippi, Hattiesburg MS

Investigators

Abstract

[unreadable] DESCRIPTION (provided by applicant): It is clear that the study of proteomics is becoming a major component of many medical, biological, and biochemical studies. It is widely accepted that proteomics studies hold much promise in the treatment and diagnosis of many diseases. With all of the promise of proteomics it is still understood that significant methodology barriers exist in performing a proteomics analysis. The proposed project herein describes novel strategies for protein modification which may aid in "top-down" protein sequencing by mass spectrometry. The hypothesis to be evaluated during this project is that gas phase free radical chemistry can be used to promote protein fragmentation within a mass spectrometer collision cell. Two aims are required to adequately test this hypothesis. The first is development of practical reagents that are capable of modifying reduced cysteine residues with free radical precursors. Our preliminary results indicate that aim 1 is possible. The second aim is to modify whole, reduced, proteins with various reagents developed in aim 1 and analyze the gas phase fragmentation chemistry which results. Our preliminary results on model compounds illustrates that predictable peptide backbone fragmentation occurs in the gas phase. We intend to prepare a variety of reagents capable of producing N-, O-, and C- centered free radicals upon activation by Collision Induced Dissociation of a weak bond in the precursor. Each of these free radicals will be studied for their ability to promote, in a selective fashion, protein backbone fragmentation via H atom abstraction/[unreadable]-fragmentation sequences. This research will provide methods for protein analysis of intact proteins which should result in increased protein sequence coverage making it possible to readily identify the important post- translational protein modifications. P This project details the development of methods to improve the analysis of whole proteins by mass spectrometry. The project has applications in the field of proteomics which is important to understanding disease. Proteomics allows for the identification of various protein levels and modifications in numerous disease states. [unreadable] [unreadable] [unreadable]