title: Conformational changes of ubiquitin during electrospray ionization as determined by in-ESI source H/D exchange combined with high-resolution MS and ECD fragmentation.
abstract: In the paper, we have demonstrated the possibility of performing hydrogen/deuterium (H/D) exchange of proteins in the region of gas-phase ion formation in an electrospray ion source by saturating the electrospray ionization source with vapors of a deuterating agent (D(2)O or MeOD). In this region, charged droplets are shrinking and the protein ions transfer into the gas phase. As a model protein, we have used ubiquitin whose ion mobility spectrometry and gas-phase H/D exchange in the vacuum part of a mass spectrometer demonstrated the presence of gas-phase conformers with different cross sections and H/D exchange rates. In our experiments, we observed monomodal deuterium distributions for all solvents, charge states, desolvating capillary temperature and types of deuterating agent. Also, we found that the number of H/D exchanges increases with an increasing desolvating capillary temperature and decreasing charge state. We observed that solution composition (49 : 50 : 1 H(2)O : MeOH : formic acid or 99 : 1 H(2)O : formic acid) influences the charge-state distribution but did not change the degree of H/D exchange for the same charge state. Electron-capture dissociation fragmentation shows that higher charge states contain a segment that is protected from access by the deuterating agent.
authors.affiliation: Russian Academy of Sciences, Institute for Energy Problems of Chemical Physics, Leninskij pr. 38 k.2, 119334, Moscow, Russia; Moscow Institute of Physics and Technology, 141700, Dolgoprudnyi, Moscow Region, Russia.
authors.forename: Yury
authors.initials: Y
authors.lastname: Kostyukevich
authors.forename: Alexey
authors.initials: A
authors.lastname: Kononikhin
authors.forename: Igor
authors.initials: I
authors.lastname: Popov
authors.forename: Eugene
authors.initials: E
authors.lastname: Nikolaev
mesh_terms: D000595:Amino Acid Sequence; D041961:Deuterium Exchange Measurement / Q000379:methods*; D005561:Formates; D000432:Methanol; D008969:Molecular Sequence Data; D011487:Protein Conformation; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013696:Temperature; D025801:Ubiquitin / Q000737:chemistry*
chemical_list: D005561:Formates; D025801:Ubiquitin; C030544:formic acid; D000432:Methanol
keywords: ECD fragmentation; FT ICR; electrospray ionization; hydrogen/deuterium exchange; in-ESI source H/D exchange; protein structure