Research Overview

Mission

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 holistic understanding of biomolecular assemblies. The Bush Lab develops and applies mass spectrometry based techniques that are fast, sensitive, and tolerant of heterogeneity for characterizing the native structures, interactions, and dynamics 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 biological 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: Theresa Gozzo
  • Degronomics: Mapping the Interacting Peptidome of a Ubiquitin Ligase Using an Integrative Mass Spectrometry Strategy. Daniele Canzani, Domnita̧ -Valeria Rusnac, Ning Zheng, and Matthew F. Bush. Anal. Chem. 2019, 91, 12775–12783. (Link)
  • 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)
  • 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 2013, 496, 64–68. (Link|PUBMED)

Small Heat Shock Proteins

Small heat shock proteins (sHSPs) are chaperones that prevent protein aggregation. sHSPs interact with clients and other sHSPs through weak, transient interactions that have resisted high-resolution structural characterization. To probe these interactions, we are developing new crosslinking MS methods, which we hypothesize will also be useful for investigating other proteins that contain less-ordered elements. We pursue this research in close collaboration with the Klevit Lab (Prof. Rachel Klevit, UW Biochemistry), which provides exciting opportunities to participate in interdisciplinary research using a variety of biophysical technique.

Crosslinking originating from a photoactive amino acid incorporated a sites 33 or 38 in the small heat shock protein HSPB5.
  • Lab Contact: Lindsey Ulmer
  • HSPB5 disease-associated mutations have long-range effects on structure and dynamics through networks of quasi-ordered interactions. Christopher N. Woods, Lindsey D. Ulmer, Maria K. Janowska, Natalie L. Stone, Ellie I. James, Miklos Guttman, Matthew F. Bush, Rachel E. Klevit. bioRxiv. , , DOI:10.1101/2022.05.30.493970. (Link)
  • Disordered Region Encodes α-crystallin Chaperone Activity Towards Lens Client γD-crystallin. Christopher N. Woods, Lindsey D. Ulmer, Miklos Guttman, Matthew F. Bush, Rachel E. Klevit. Proc. Natl. Acad. Sci. 2023, 120, e221376512. (Link)

Innovations in Instrumentation

The Bush Lab is focused on pushing the frontiers of chemical measurements, including maximizing the value of measurements using existing commercial instruments and developing next-generation instruments with unique functionality. We’re particularly interested in standardizing ion mobility measurements and increasing the orthogonality between ion mobility and mass spectrometry. Towards that end, we’ve developed radio-frequency confining drift cells, which we use for determining absolute collision cross section values, and extended the Structures for Lossless Ion Manipulations (SLIM) architecture to native-like ions, which we use for multidimensional ion mobility. Members of the Bush Lab benefit from the resources of the Student Innovation Center.

Our latest platform for tandem ion mobility, mass spectrometry.
  • Lab Contact: AnneClaire Wageman
  • A Flexible, Modular Platform for Multidimensional Ion Mobility of Native-like Ions. Rachel M. Eaton, Benjamin Zercher, AnneClaire Wageman, Matthew F. Bush. J. Am. Soc. Mass Spectrom. 2023, 34, 1175–1185. (Link)
  • Analysis of Native-Like Ions using Structures for Lossless Ion Manipulations. Samuel J. Allen, Rachel M. Eaton, Matthew F. Bush. Anal. Chem. 2016, 88, 9118–9126. (Link)

Relationship between Structure, Charge State, and Collision Cross Section

Ion mobility mass spectrometry probes the structures of protein ions in the gas phase, but electrospray ionization yields ions with a range of charge state and ions of different charge state usually exhibit different collision cross section values. No consensus has emerged regarding how results for different charge states should be integrated into the structural elucidation process. The Bush Lab uses experimental and computational methods to probe the relationship between these properties with the objective of understanding the relationship between the structures of proteins in solution and the observables of structural MS experiments.

  • Lab Contact: Theresa Gozzo
  • Effects of charge on protein ion structure: Lessons from cation-to-anion, proton-transfer reactions. Theresa A. Gozzo, Matthew F. Bush. Mass Spectrom. Reviews 2023, , DOI:10.1002/mas.21847. (Link)
  • Folding of Protein Ions in the Gas Phase after Cation-to-Anion Proton-Transfer Reactions (CAPTR). Kenneth J. Laszlo, Eleanor B. Munger, and Matthew F. Bush. J. Am. Chem. Soc. 2016, 138, 9581–9588. (Link|PUBMED)
  • 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|PUBMED)

Ion Mobility Theory

The Bush Lab develops theory to predict the collision cross sections of ions based on atomic models and the information content of ion mobility separations based on the design of the experiment. The outcomes of these projects increase the value of current ion mobility measurements and guide our efforts to develop higher-performance implementations of ion mobility.

Interaction potential between a gas molecule and a ubiquitin ion.
  • Lab Contacts: Theresa Gozzo
  • Ion Mobility of Proteins in Nitrogen Gas: Effects of Charge State, Charge Distribution, and Structure. Daniele Canzani, Kenneth J. Laszlo, Matthew F. Bush. J. Phys. Chem. A 2018, 122, 5625−5634. (Link)
  • Effects of Drift Gas Selection on the Ambient-Temperature, Ion Mobility Mass Spectrometry Analysis of Amino Acids. Kimberly L. Davidson and Matthew F. Bush. Anal. Chem. 2017, 89, 2017–2023. (Link)