Abstract
The folding pathways of gas-phase cytochrome c ions produced by electrospray ionization have been studied by an ion trapping/ion mobility technique that allows conformations to be examined over extended timescales (10 ms to 10 s). The results show that the +9 charge state emerges from solution as a compact structure and then rapidly unfolds into several substantially more open structures, a transition that requires 30–60 ms; over substantially longer timescales (250 ms to 10 s) elongated states appear to refold into an array of folded structures. The new folded states are less compact than those that are apparent during the initial unfolding. Apparently, unfolding to highly open conformations is a key step that must occur before +9 ions can sample more compact states that are stable at longer times.
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Bai, Y. W.; Sosnick, T. R.; Mayne, L.; Englander, S. W. Protein Folding Intermediates: Native-State Hydrogen Exchange. Science 1995, 269, 192–197.
Dobson, C. M.; Sali, A.; Karplus, M. Protein Folding: A Perspective From Theory and Experiment. Angew. Chem. Int. Ed. Engl. 1998, 37, 868–893.
Gruebele, M. The Fast Protein Folding Problem. Annu. Rev. Phys. Chem. 1999, 50, 485–516.
Shelimov, K. B.; Clemmer, D. E.; Hudgins, R. R.; Jarrold, M. F. Protein Structure in Vacuo: The Gas Phase Conformations of BPTI and Cytochrome c. J. Am. Chem. Soc. 1997, 119, 2240–2248.
Wyttenbach, T.; Helden, G. V.; Bowers, M. T. Gas-Phase. Conformation of Biological Molecules: Bradykinin. J. Am. Chem. Soc. 1996, 118, 8355–8364.
Valentine, S. J.; Clemmer, D. E. H/D Exchange Levels of Shape-Resolved Cytochrome c Conformers in the Gas Phase. J. Am. Chem. Soc. 1997, 119, 3558–3566.
Winger, B. E.; Lightwahl, K. J.; Rockwood, A. L.; Smith, R. D. Probing Qualitative Conformation Differences of Multiply Protonated Gas-Phase Proteins via Hydrogen/Deuterium Isotopic Exchange With Water-d2. J. Am. Chem. Soc. 1992, 114, 5897–5898.
Wood, T. D.; Chorush, R. A.; Wampler, F. M.; Little, D. P.; Oconnor, P. B.; McLafferty, F. W. Gas-Phase Folding and Unfolding of Cytochrome c Cations. Proc. Natl. Acad. Sci. U. S. A. 1995, 92, 2451–2454.
McLafferty, F. W.; Guan, Z. Q.; Haupts, U.; Wood, T. D.; Kelleher, N. L. Gaseous Conformational Structures of Cytochrome. c. J. Am. Chem. Soc. 1998, 120, 4732–4740.
Freitas, M. A.; Hendrickson, C. L.; Emmett, M. R.; Marshall, A. G. Gas-Phase Bovine Ubiquitin Cation Conformations Resolved by Gas-Phase Hydrogen/Deuterium Exchange Rate and Extent. Int. J. Mass Spectrom. 1999, 187, 565–575.
Horn, D. M.; Breuker, K.; Frank, A. J.; McLafferty, F. W. Kinetic Intermediates in the Folding of Gaseous Protein Ions Characterized by Electron Capture Dissociation Mass Spectrometry. J. Am. Chem. Soc. 2001, 123, 9792–9799.
Breuker, K.; Oh, H. B.; Horn, D. M.; Cerda, B. A.; McLafferty, F. W. Detailed Unfolding and Folding of Gaseous Ubiquitin Ions Characterized by Electron Capture Dissociation. J. Am. Chem. Soc. 2002, 124, 6407–6420.
Zubarev, R. A.; Horn, D. M.; Fridriksson, E. K.; Kelleher, N. L.; Kruger, N. A.; Lewis, M. A.; Carpenter, B. K.; McLafferty, F. W. Electron Capture Dissociation for Structural Characterization of Multiply Charged Protein Cations. Anal. Chem. 2000, 72, 563–573.
Woenckhaus, J.; Mao, Y.; Jarrold, M. F. Hydration of Gas Phase Proteins: Folded +5 and Unfolded +7 Charge States of Cytochrome. c. J. Phys. Chem. B 1997, 101, 847–851.
Jarrold, M. F. Peptides and Proteins in the Vapor Phase. Annu. Rev. Phys. Chem. 2000, 51, 179–207.
Woenckhaus, J. Drift Time Mass Spectrometric Protein Hydration Experiments. Int. J. Mass Spectrom. 2002, 213, 9–24.
Badman, E. R.; Hoaglund-Hyzer, C. S.; Clemmer, D. E. Monitoring Structural Changes of Proteins in an Ion Trap over ∼10–200 ms: Unfolding Transitions in Cytochrome c Ions. Anal. Chem. 2001, 73, 6000–6007.
Myung, S.; Badman, E. R.; Lee, Y. J.; Clemmer, D. E. Structural Transitions of Electrosprayed Ubiquitin Ions Stored in an Ion Trap over ∼10 ms to 30 s. J. Phys. Chem. A. 2002, 106, 9976–9982.
Hoaglund-Hyzer, C. S.; Li, J. W.; Clemmer, D. E. Mobility Labeling for Parallel CID of Ion Mixtures. Anal. Chem. 2000, 72, 2737–2740.
Clemmer, D. E.; Jarrold, M. F. Ion Mobility Measurements and Their Applications to Clusters and Biomolecules. J. Mass Spectrom. 1997, 32, 577–592.
Valentine, S. J.; Anderson, J. G.; Ellington, A. D.; Clemmer, D. E. Disulfide-Intact and -Reduced Lysozyme in the Gas Phase: Conformations and Pathways of Folding and Unfolding. J. Phys. Chem. B 1997, 101, 3891–3900.
McLuckey, S. A.; Wells, J. M.; Stephenson, J. L.; Goeringer, D. E. Novel Quadrupole Ion Trap Methods for Characterizing the Chemistry of Gaseous Macro-Ions. Int. J. Mass Spectrom. 2000, 200, 137–161.
Badman, E. R.; Myung, S.; Clemmer, D. E. Gas-Phase Separations of Protein and Peptide Ion Fragments Generated by Collision-Induced Dissociation in an Ion Trap. Anal. Chem. 2002, 74, 4889–4894.
Shvartsburg, A. A.; Jarrold, M. F. An Exact Hard-Spheres Scattering Model for the Mobilities of Polyatomic Ions. Chem. Phys. Lett. 1996, 261, 86–91.
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Published online July 12, 2005
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Badman, E.R., Myung, S. & Clemmer, D.E. Evidence for unfolding and refolding of gas-phase cytochrome c ions in a Paul trap. J Am Soc Mass Spectrom 16, 1493–1497 (2005). https://doi.org/10.1016/j.jasms.2005.04.013
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DOI: https://doi.org/10.1016/j.jasms.2005.04.013