Evaluation of a Commercial TIMS-Q-TOF Platform for Native Mass Spectrometry

17 April 2024, Version 3
This content is a preprint and has not undergone peer review at the time of posting.

Abstract

Mass-spectrometry based assays in structural biology studies measure either intact or digested proteins. Typically, different mass spectrometers are dedicated for such measurements: those optimized for rapid analysis of peptides or those designed for high molecular weight analysis. A commercial trapped ion mobility-quadrupole-time of flight (TIMS-Q-TOF) platform is widely utilized for proteomics and metabolomics, with ion mobility providing a separation dimension in addition to liquid chromatography. The ability to perform high-quality native mass spectrometry of protein complexes, however, remains largely uninvestigated. Here, we evaluate a commercial TIMS-Q-TOF platform for analyzing non-covalent protein complexes by utilizing the instrument’s full range of ion mobility, MS, and MS/MS (both in-source activation and collision cell CID) capabilities. The TIMS analyzer is able to be tuned gently to yield collision cross sections on native-like complexes comparable to those previously reported on various instrument platforms. In-source activation and collision cell CID were robust for both small and large complexes. TIMS-CID was performed on protein complexes streptavidin (53 kDa), avidin (68 kDa), and cholera toxin B (CTB, 58 kDa). Complexes pyruvate kinase (237 kDa) and GroEL (801 kDa) were beyond the trapping capabilities of the commercial TIMS analyzer, but mass spectra could be acquired. The presented results indicate that the commercial TIMS-Q-TOF platform can be used for both omics and native mass spectrometry applications; however, modifications to the commercial RF drivers for both the TIMS analyzer and quadrupole (currently limited to m/z 3,000) are necessary to mobility analyze protein complexes greater than about 60 kDa.

Keywords

Native Mass Spectrometry
Ion Mobility
TIMS
Protein Complexes

Supplementary materials

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Title
Supporting Information: Evaluation of a Commercial TIMS-Q-TOF Platform for Native Mass Spectrometry
Description
The supplemental information contains information about the samples analyzed in this work, a comparison of typical proteomics tune settings and those for native MS experiments, comparison of sample concentration on mobility, comparison of TIMS duty cycle on mobility, mass spectra with raw intensities.
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