Research

Most proteins, particularly those that accomplish complicated tasks, form assemblies with other proteins and molecules that are critical to their function. Established structural biology tools are most effective for highly purified samples that have limited conformational variability, which makes it challenging to apply those methods to capture a systems-wide understanding of the structures, interactions, and dynamics that are present under different cellular conditions. The Bush Lab develops and applies mass spectrometry based techniques that are fast, sensitive, and tolerant of heterogeneity for characterizing the native structures of biological assemblies.


Modelling biomolecular assemblies using constraints from mass spectrometry (MS) and ion mobility (IM) experiments. The masses and identities of individual proteins (subunits) are determined using proteomics (a). The stoichiometry of the intact assembly is determined using MS from a native-like buffer (b). Subassemblies are generated by disrupting the assembly in solution and characterized by MS (c). Collision cross sections (Ω) of the intact assembly and subassemblies provide conformational information (d). The masses of subunits and subassemblies from MS are used to generate 2D interaction maps (e). 3D models are constructed using these maps, Ω values, and any complementary structural information (f). Atomic structures and models can then be docked into these models (g).

Native Mass Spectrometry. Gas-phase ions of assemblies can retain significant memories of their native structures in solution. Many measurements of stoichiometry, connectivity, and shape have shown that these aspects of assembly structure can be strongly correlated in both environments. Gas-phase techniques, including mass spectrometry, ion mobility, and ion chemistry are used to probe the native structures of biological assemblies. Accurate models of protein assemblies, including those that are heterogeneous, dynamic, and/or membrane-bound, are built based on results from both gas-phase experiments and complementary techniques.

  • Lab Contacts: Sam Marionni & Tracy Stanzel
  • Hexamers of the Type II Secretion ATPase GspE from Vibrio cholerae with Increased ATPase Activity Connie Lu, Stewart Turley, Samuel T. Marionni, Young-Jun Park, Kelly K. Lee, Marcella Patrick, Ripal Shah, Maria Sandkvist, Matthew F. Bush, Wim G.J. Hol. Structure 2013, 21, 1707–1717. (Link|PUBMED)
  • Ion Mobility Mass Spectrometry of a Circadian Clock Protein Complex Reveals a Ligand-Dependent Conformational Switch Samuel T. Marionni, Weiman Xing, Ning Zheng, Matthew F. Bush. 60th American Society for Mass Spectrometry Conference 2012 (Poster). SCFFBXL3 ubiquitin ligase targets cryptochromes at their cofactor pocket Weiman Xing, Luca Busino, Thomas R. Hinds, Samuel T. Marionni, Nabiha H. Saifee, Matthew F. Bush, Michele Pagano, Ning Zheng. Nature 2013496, 64–68. (Link|PUBMED)

toc_polarity_120-01

Electrospray Fundamentals. In native mass spectrometry, intact protein complexes are transferred from solution into the gas phase using nanoelectrospray ionization. We use a range of mass spectrometry and ion mobility experiments, coupled with computational chemistry and Monte Carlo simulations, to gain fundamental insights into our implementations of nanoelectrospray ionization. These projects probe many stages of this process, from the initial nanodroplets to the final multiply charged ions. The outcomes of these projects are guiding our long term objectives of increasing the sensitivity of native mass spectrometry experiments and the information content of the resulting native mass spectra.

  • Lab Contacts: Sam Allen & Kim Davidson
  • Effects of Polarity on the Structures and Charge States of Native-like Proteins and Protein Complexes in the Gas Phase Samuel J. Allen, Alicia M. Schwartz, Matthew F. Bush. Anal. Chem. 2013, 85, 12055–12061. (Link)

RF Confining Drift Cell

A new RF confining drift cell developed for use on our Waters Synapt G2 HDMS. We use this cell to measure absolute collision cross sections for ions of intact protein complexes.

Ion Mobility (IM) Mass Spectrometry (MS) Development. IM is an emerging technology for characterizing the structures of biomolecules that can be used in tandem with MS. IM separates ions based primarily on their charge state and collision cross section, which depends on ion shape and other factors. We are developing new IM cells to enable the measurement of high accuracy collision cross sections and new methods for interpreting those results, in the contexts of both native MS and systems biology.

  • Lab Contact: Sam Allen
  • Design and Characterization of a New Ion Mobility Cell for Protein Complexes Samuel J. Allen, Samuel T. Marionni, Kevin Giles, Tony Gilbert, Matthew F. Bush. 60th American Society for Mass Spectrometry Conference 2012 (Poster)
  • Ion mobility mass spectrometry of peptide ions: effects of drift gas and calibration strategies Matthew F. Bush, Iain D. G. Campuzano, Carol V. Robinson. Anal. Chem. 201284, 7124–7130. (Link|PUBMED)
  • Structural Characterization of Drug-like Compounds by Ion Mobility Mass Spectrometry: Comparison of Theoretical and Experimentally Derived Nitrogen Collision Cross-sections Iain Campuzano, Matthew F. Bush, Carol V. Robinson, Claire Beaumont, Keith Richardson, Hyungjun Kim, Hugh I. Kim. Anal. Chem. 201284, 1026-1033. (Link|PUBMED)

Ion Mobility Arrival Time Distribution of (AAHAL+2H)^2+

The ion mobility arrival time distribution of doubly protonated AAHAL is sensitive to subtle details of ion structure.

Structures of Peptide Ions & their Dissociation Products. In collaboration with the Turecek group, we are investigating the structures of peptide ions and their gas-phase dissociation products. Ions are activated using either collision-induced dissociation or electron-transfer dissociation prior to characterization using ion mobility. These studies provide fundamental insights into the structures and reactivities of peptide ions, which can increase the information content of  proteomics experiments.

  • Lab Contact: Ken Laszlo
  • Comprehensive Analysis of Gly-Leu-Gly-Gly-Lys Peptide Dication Structures and Cation-Radical Dissociations Following Electron Transfer: From Electron Attachment to Backbone Cleavage, Ion-Molecule Complexes, and Fragment Separation Robert Pepin, Kenneth J. Laszlo, Bo Peng, Aleš Marek, Matthew F. Bush, František Tureček. J. Phys. Chem. A 2014118, 308–324. (Link|PUBMED)
  • Gas-Phase Structures of Phosphopeptide Ions: A Difficult Case František Tureček, Christopher L. Moss, Ioannis Pikalov, Robert Pepin, Kerim Gulyuz, Nicolas C. Polfer, Matthew F. Bush, Jeffery Brown, Jonathan Williams, Keith Richardson. Int. J. Mass Spectrom. 2013354–355, 249–256. (Link)
  • Electron Transfer Dissociation of Photolabeled Peptides. Backbone Cleavages Compete with Diazirine Ring Rearrangements Aleš Marek, Robert Pepin, Bo Peng, Kenneth J. Laszlo, Matthew F. Bush, František Tureček. J. Am. Soc. Mass Spectrom. 201324, 1641–1653. (Link|PUBMED)
  • Assigning Structures to Gas-Phase Peptide Cations and Cation-Radicals. An Infrared Multiphoton Dissociation, Ion Mobility, Electron Transfer, and Computational Study of a Histidine Peptide Ion Christopher L. Moss, Julia Chamot-Rooke, Edith Nicol, Jeffery Brown, Iain Campuzano, Keith Richardson, Jonathan P. Williams, Matthew F. Bush, Benjamin Bythell, Bela Paizs, Frantisek Turecek. J. Phys. Chem B 2012116, 3445–3456. (Link|PUBMED)

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