Research Overview

The Wysocki group is divided into three subgroups, Instrumentation, Complexes, and Omics. (1) Instrumentation is involved in native MS fundamentals, method development and modification including design and implementation of surface-induced dissociation devices, collision cross-section measurements with ion mobility, UV photodissociation of proteins and protein complexes, and separations coupled to native MS. (2) Complexes is involved in collaborations with a focus on the application of native MS for the structural characterization of protein-protein, protein-ligand, protein-RNA, and protein-DNA interactions. (3) Omics includes traditional MS-based proteomics and metabolomics studies for the detection and treatment of infectious and noninfectious diseases. Research can be categorized into the following four broad areas for which members from any subgroup might participate:

(1) Surface-Induced Dissociation Development and Implementation

Subunit organization of protein complexes can be studied with tandem mass spectrometry by disruption of the quaternary structure in a controlled manner. The major challenge of using this method for structural analysis of non-covalent protein complexes is to overcome the undesired unfolding of subunits that occurs in the commonly used collision activation with gaseous targets, resulting in loss of original structural information. This area of research involves the development of surface-induced dissociation (SID) devices for improved structural characterization of large protein complexes. We aim to create devices that are “vendor-neutral” and allow for straightforward operation by the end-user. Our custom SID devices reveal protein complex architecture, including subunit stoichiometry and connectivity, and minimal or reduced unfolding compared with CID.

(2) Native MS of Protein and Nucleoprotein Complexes

Our NIH-funded Resource for Native MS-Guided Structural Biology develops MS and related technology (e.g., online separations and ion mobility) to build an integrated structural biology approach. Collaborations with scientists from around the world provide biological problems that drive the technology development while providing group members with collaborative science training and expertise in handling and characterization of protein, nucleoprotein, and membrane protein complexes.

(3) Proteomics and Metabolomics

A third area of research in the Wysocki lab is combined proteomics and metabolomics analysis to study different disease systems. This area of research ranges from understanding microbial ecology during Salmonella infection to decoding deregulated pathways in lung cancer. Expansion of the Salmonella project includes characterizing metabolic exchanges between Salmonella and its competitors in the gut as well as protein profiling to identify nutrient acquisition and utilization pathways that could correspond to consumption of metabolites by the microbiota. This research will facilitate improved prophylactic and therapeutic options in the future.

(4) Fragment Ion Structures by IRMPD

These projects are designed to increase the current understanding of the fragmentation patterns of activated protonated peptides and other ions (glycans, RNA/DNA). The long-range goals of this work are to provide additional “rules” that will, ultimately, relate information on gas-phase fragmentation patterns and energetics of dissociation to the gas-phase conformations of intact and fragmented molecules. The investigation of fragmentation mechanisms/product ion structure is primarily performed by combining experimental data with computationally derived structures and their theoretical infrared spectra. Experimental data are collected using free electron laser (FEL) Infrared Multiphoton Dissociation (IRMPD) spectroscopy in which a mass spectrometer is used to detect fragments as a function of infrared wavelength, generating an action IR spectrum.