Ultraviolet Photodissociation Mass Spectrometry for Characterization of Biological Molecules

Abstract. Understanding the functions of lipids, proteins and even larger macromolecular assemblies
depends on deciphering complex structures of individual molecules as well as decrypting how those
molecules interact, often via networks of non-covalent interactions. In order to advance the elucidation of
biomolecular organization and functional outcomes, new methods are needed to push the limits of
structural insight, providing more detailed holistic chemical information with greater sensitivity. The critical
interplay between structure/function is evidenced in numerous biologically-motivated problems, ranging
from understanding the ways that pathogenic bacteria develop antibiotic resistance to the design of new
drugs that selectively bind and inhibit the functions of protein targets. The ongoing need for even greater
chemical insight has motivated my group’s effort to develop innovative mass spectrometry methods to
characterize structures of biological molecules in unprecedented detail, especially lipids and proteins
which are featured in this proposal. The overarching goal of my research program is to develop state-of-
the-art tandem mass spectrometry technologies, particularly highlighting ultraviolet photodissociation
(UVPD) and hybrid MS/MS methods, for structural elucidation of lipids, proteins, and protein complexes.
These new methods will be showcased for solving challenging problems in three areas. (1) Lipids: (i)
profiling lipids of pathogenic bacteria and their signatures of antibiotic resistance, and (ii) structural
characterization of unsaturations, oxidations and other modifications of lipids that occur during
remodeling of cellular membranes. (2) Protein complexes: (i) characterization of protein-ligand
complexes, membrane protein complexes, protein/nucleic acid complexes, and macromolecular
assemblies, and (ii) advancing capillary electrophoresis for native separations and exploration of the
interactome. (3) Post-translational modifications: focusing on decoding the phosphorylation patterns of
the C-terminal domain of RNA polymerase II which regulates transcription. These high impact problems
are supported via numerous collaborations with microbiology and molecular biology groups who
recognize the value of frontier mass spectrometry strategies for elevating biomedical research.

Grant Number: 5R35GM139658-05
NIH Institute/Center: NIH
Principal Investigator: Jennifer Brodbelt