Journal
PROTEOMICS
Volume 15, Issue 16, Pages 2835-2841Publisher
WILEY
DOI: 10.1002/pmic.201400457
Keywords
Animal proteomics; Electrospray ionization; Ion mobility-mass spectrometry; Protein conformation; Ubiquitin specific protease 5
Funding
- University of Nottingham
- Biotechnology and Biological Sciences Research Council [BB/F019297/1] Funding Source: researchfish
- BBSRC [BB/F019297/1] Funding Source: UKRI
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Many proteins exhibit conformation flexibility as part of their biological function, whether through the presence of a series of well-defined states or by the existence of intrinsic disorder. Ion mobility spectrometry, in combination with MS (IM-MS), offers a rapid and sensitive means of probing ensembles of protein structures through measurement of gas-phase collisional cross sections. We have applied IM-MS analysis to the multidomain deubiquitinating enzyme ubiquitin specific protease 5 (USP5), which is believed to exhibit significant conformational flexibility. Native ESI-MS measurement of the 94-kDa USP5 revealed two distinct charge-state distributions: [M + 17H](+) to [M + 21H](+) and [M + 24H](+) to [M + 29H](+). The collisional cross sections of these ions revealed clear groupings of 52 +/- 4 nm(2) for the lower charges and 66 +/- 6nm(2) for the higher charges. Molecular dynamics simulation of a compact form of USP5, based on a crystal structure, produced structures of 53-54 nm(2) following 2 ns in the gas phase, while simulation of an extended form (based on small-angle X-ray scattering data) led to structures of 64 nm(2). These data demonstrate that IM-MS is a valuable tool in studying proteins with different discrete conformational states.
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