title: Ion Mobility-Mass Spectrometry Analysis of Cross-Linked Intact Multiprotein Complexes: Enhanced Gas-Phase Stabilities and Altered Dissociation Pathways.
abstract: Analysis of protein complexes by ion mobility-mass spectrometry is a valuable method for the rapid assessment of complex composition, binding stoichiometries, and structures. However, capturing labile, unknown protein assemblies directly from cells remains a challenge for the technology. Furthermore, ion mobility-mass spectrometry measurements of complexes, subcomplexes, and subunits are necessary to build complete models of intact assemblies, and such data can be difficult to acquire in a comprehensive fashion. Here, we present the use of novel mass spectrometry cleavable cross-linkers and tags to stabilize intact protein complexes for ion mobility-mass spectrometry. Our data reveal that tags and linkers bearing permanent charges are superior stabilizers relative to neutral cross-linkers, especially in the context of retaining compact forms of the assembly under a wide array of activating conditions. In addition, when cross-linked protein complexes are collisionally activated in the gas phase, a larger proportion of the product ions produced are often more compact and reflect native protein subcomplexes when compared with unmodified complexes activated in the same fashion, greatly enabling applications in structural biology.
authors.affiliation: Departments of †Chemistry, ‡Biological Chemistry, and §Computational Medicine and Bioinformatics, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Billy M
authors.initials: BM
authors.lastname: Samulak
authors.affiliation: Departments of †Chemistry, ‡Biological Chemistry, and §Computational Medicine and Bioinformatics, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Shuai
authors.initials: S
authors.lastname: Niu
authors.affiliation: Departments of †Chemistry, ‡Biological Chemistry, and §Computational Medicine and Bioinformatics, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Philip C
authors.initials: PC
authors.lastname: Andrews
authors.affiliation: Departments of †Chemistry, ‡Biological Chemistry, and §Computational Medicine and Bioinformatics, University of Michigan , Ann Arbor, Michigan 48109, United States.
authors.forename: Brandon T
authors.initials: BT
authors.lastname: Ruotolo
mesh_terms: D000818:Animals; D005740:Gases / Q000737:chemistry; D000075663:Ion Mobility Spectrometry; D007477:Ions / Q000737:chemistry; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000378:metabolism; D055550:Protein Stability; D020836:Protein Structure, Quaternary; D014305:Triose-Phosphate Isomerase / Q000737:chemistry / Q000378:metabolism
chemical_list: D005740:Gases; D007477:Ions; D046912:Multiprotein Complexes; D014305:Triose-Phosphate Isomerase