title: Gas-phase protein conformation/multimer ion formation by electrospray ion mobility-mass spectrometry: bovine insulin and ubiquitin.
abstract: Ion mobility-mass spectrometry (IMMS) is a very attractive method for studies in structural biology because of the ability of rapid isolation by nearly simultaneous m/z characterization and size separation, leading to an emergence of IMMS as a complimentary biochemical tool. Earlier, we developed a method based on varying the protein concentration in solution prior to electrospray ionization (ESI) with subsequent m/z selection and dissociation of protein multimers by IMMS of cytochrome c. The focus of this work will be to correctly distinguish truly different ion conformations formed by ESI versus homomultimeric complexes with the same m/z for well-studied proteins bovine ubiquitin and insulin. These proteins were chosen due to their large difference in solution phase structures: insulin tightly bound by disulfide linkages, and ubiquitin-a protein that may adopt a range of states from compact to extended. Our preliminary results, as with cytochrome c reveal false negatives for protein oligomer formation and false positives for protein conformational states. In addition, these results will be couched in terms of the need for quantification of IMMS analysis of proteins given the total area under IMMS peaks can also distinguish conformation versus aggregation as higher order oligomers have more mass per ion.This article is part of the themed issue 'Quantitative mass spectrometry'.
authors.affiliation: Genomics Research Center, Academia Sinica, 128 Academia Road, Nangang Section 2, Taipei 115, Taiwan, Republic of China kgillig@gate.sinica.edu.tw.
authors.forename: Kent J
authors.identifier: http://orcid.org/0000-0002-4937-1940
authors.initials: KJ
authors.lastname: Gillig
mesh_terms: D000818:Animals; D002417:Cattle; D005740:Gases / Q000737:chemistry*; D061385:Insulins / Q000737:chemistry*; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D021241:Spectrometry, Mass, Electrospray Ionization; D025801:Ubiquitin / Q000737:chemistry*
chemical_list: D005740:Gases; D061385:Insulins; D025801:Ubiquitin
keywords: aggregation; ion mobility-mass spectrometry; protein structure; quantification