CAREER: Developing hybrid IMS/OMS-MS/MS techniques for protein complex structure characterization

West Virginia University Research Corporation, Morgantown WV

Investigators

Abstract

With support from the Chemical Measurement and Imaging Program in the Division of Chemistry and the Instrument Development for Biological Research (IDBR) Program in the Division of Biological Infrastructure, Professor Valentine at West Virginia University and his group are developing scientific instrumentation and techniques for studying the three-dimensional shapes of complex, biological molecules. This developmental work is aimed at the study of protein complexes. Proteins play a crucial role in life processes. When such molecules do not form the correct three dimensional shapes, disease often ensues and therefore it becomes important to be able to determine the structures of these biological molecules and how their components come to form critical shapes. However, because of their relatively large sizes and other factors, it is difficult to determine accurate structures for many molecules. The new instrumentation and techniques developed by Professor Valentine are important ways for determining the structures of such proteins. Professor Valentine is also engaged in educational outreach efforts to improve the training of Middle- and High-School teachers in the science, technology, engineering and mathematical fields thereby benefitting many students. The characterization of protein complex structures as well as the structures of coexisting protein species presents one challenge to modern analytical techniques. Professor Valentine is developing novel gas-phase separation instrumentation and techniques to study these problems. This work has three major tasks. First, a hybrid drift tube that combines traditional ion mobility spectrometry (IMS) separations with overtone mobility spectrometry (OMS) analysis is constructed and coupled with an existing linear ion trap mass spectrometer. The approach may allow the probing of minor conformational changes (~4%) occurring on very short timescales (microsecond and lower), enhancing current methods for structural assessments. Second, the novel technique is used to characterize a protein system associated with Huntington's Disease. Multimeric peptide ions formed by portions of the huntingtin (Htt) protein are subjected to OMS-hydrogen deuterium exchange (HDX) analysis with tandem mass spectrometry (MS). The results are compared with those obtained from IMS-OMS-MS analysis to provide a means for testing the new instrumentation and techniques on a real-world sample as well as provide new information in the form of species that are undergoing structural transformations during the course of the measurement. The final task is characterizing these same protein systems using solution-phase HDX combined with OMS-MS/MS studies to unravel linking of solution- and gas-phase structures as well as to provide information regarding conformation establishment in the gas-phase. This research may provide important clues about Huntington's Disease progression. The educational component establishes strategies for improving the efficacy of middle- and high-school science instruction, develops undergraduate curricula, introduces undergraduate students to research, and provides unique research and instructional opportunities for graduate students.