Shifts in protein charge state distributions with varying redox reagents in nanoelectrospray triple quadrupole mass spectrometry

Zhao, Cheng; Wood, Troy D.; Bruckenstein, Stanley

doi:10.1016/j.jasms.2004.12.003

Cited by 12 publications

(9 citation statements)

References 55 publications

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“…Most probably as the result of occurring proton transfer reactions, increasing amounts of ascorbic acid slightly shifted peak intensity from the +2 charge state to the +1 charge state. This observation was consistent with the effect of ascorbic acid on the charge state distribution of proteins in ESI-MS [40, 41]. …”

Section: Resultssupporting

confidence: 88%

Ascorbic acid for homogenous redox buffering in electrospray ionization–mass spectrometry

Plattner

Erb

Chervet³

et al. 2012

Anal Bioanal Chem

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Electrospray ionization (ESI) involves the dispersion of a liquid containing analytes of interest into a fine aerosol by applying a high potential difference to the sample solution with respect to a counter electrode. Thus, from the electrochemical point of view, the ESI source represents a two-electrode controlled-current electrochemical flow cell. The electroactive compounds part of the solvent sprayed may be altered by occurring electrolysis (oxidation in positive ion mode and reduction in negative ion mode). These reactions can be troublesome in the context of unknown identification and quantification. In the search for a simple, inexpensive, and efficient way to suppress electrochemical oxidation in positive ESI, the usability of ascorbic acid, hydroquinone, and glutathione for homogenous redox buffering was tested. Performance of the antioxidants was assessed by analyzing pharmaceutical compounds covering a broad range of functional groups prone to oxidation. Different emitter setups were applied for continuous infusion, flow injection, and liquid chromatography/mass spectrometry experiments. Best performance was obtained with ascorbic acid. In comparison to hydroquinone and glutathione, ascorbic acid offered superior antioxidant activity, a relatively inert oxidation product, and hardly any negative effect on the ionization efficiency of analytes. Furthermore, ascorbic acid suppressed the formation of sodiated forms and was able to induce charge state reduction. Only in the very special case of analyzing a compound isobaric to ascorbic acid, interference with the low-abundant [ascorbic acid+H]+ signal may become a point of attention.FigureAscorbic acid efficiently suppresses analyte oxidation and formation of sodiated forms in positive electrospray ionizationElectronic supplementary materialThe online version of this article (doi:10.1007/s00216-012-6196-z) contains supplementary material, which is available to authorized users.

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Section: Resultssupporting

confidence: 88%

Ascorbic acid for homogenous redox buffering in electrospray ionization–mass spectrometry

Plattner

Erb

Chervet³

et al. 2012

Anal Bioanal Chem

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“…2 Indeed, use of a reducing reagent admixed with protein analyte quantifiably reduces the weighted averaged charge state (ACS) 3 distribution. For example, the reducing agent 1,4-benzoquinone shifted the multiple charge state distribution of cytochrome c from ACS D 14.25 (in the absence of 1,4-benzoquinone) to ACS D 7.10.…”

Section: Introductionmentioning

confidence: 99%

“…2 However, for the reagents hydroquinone (ACS D 14.04), ascorbic acid (ACS D 11.82), and 7,7,8,8-tetracyanoquinodimethane (TCNQ, ACS D 12.95), smaller effects were noted on the ACS of cytochrome c. These were consistent with their redox properties in that hydroquinone and ascorbic acid are reduced forms that are unlikely to accept protons, while TCNQ is an electron acceptor and thus would not be expected to assist charge reduction for multiply charged protein ions. 2 In addition, unfolding of the protein to enable more exposed sites to be involved in charge transfer was ruled out as contributing significantly to the charge reduction mechanism; far UV circular dichroism (CD) spectra of cytochrome c and ubiquitin did not vary significantly upon the admixing of these proteins with any of the redox reagents, indicating that conformation is affected to a small extent (less than 10% loss in ellipticity) by the presence of the redox reagents tested. Furthermore, the observed ACS reduction did not correlate with the gas-phase basicities of the reagents, i.e.…”

Section: Introductionmentioning

confidence: 99%

“…It was not clear from our earlier data whether the mechanism of ACS reduction was a result of proton-transfer reactions and/or electron-transfer reactions. To resolve this ambiguity, 2 we employed ESI-FT-ICR mass spectrometry, which is well-known for its powerful ability to conduct exact mass measurements with very low reported mass measurement error(s) 7 -11 and to provide isotopically resolved charge state distributions from proteins up to 112 kDa. 12 Johnson et al have shown previously the power of ESI-FT-ICR to observe isotopic distributions of metalloproteins to distinguish between oxidized and reduced states of the metal.…”

Section: Introductionmentioning

confidence: 99%

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FT‐ICRMS distinguishes the mechanism of the charge state reduction for multiply charged protein cations admixed with redox reagents in ESI

et al. 2006

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Recently, we reported on a phenomenon in which multiply charged protein cations produced by electrospray ionization could be reduced to lower and narrower charge state distributions when admixed with reducing reagents 1,4-benzoquinone or quinhydrone. Circular dichroism spectra of the proteins indicated that secondary and tertiary structural changes upon addition of these reducing reagents were negligible, thus eliminating conformational effects as playing a role in the charge reduction mechanism. Furthermore, the extent of charge state reduction did not correspond with gas-phase basicities of the redox reagents, suggesting that solution-phase, and not gas-phase, behavior dominates the observed charge state reduction. The relatively low resolution of the triple quadrupole employed did not make it possible to distinguish isotopic distributions of the multiply charged cations in order to determine whether the observed phenomenon was the result of proton-transfer reactions between the multiply charged cations and the reducing reagent or because of electron transfer from the reducing reagent to the protein cations. Here, high-resolution ESI-Fourier transform ion cyclotron resonance mass spectrometry of several peptide amides in the presence of a redox reagent show isotopic distributions that are consistent only with the proton-transfer mechanism.

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“…One strategy is to manipulate solution conditions, such as solvent composition and pH [10]. Other approaches involve the addition of "super-charging" [13,[17][18][19][20][21] or redox agents [22]. Methods for the control of the charge state distribution by gas-phase proton transfer via ion/ion [23,24] and ion/ molecular [25][26][27] reactions have also been reported, which eliminates many of the drawbacks of adding external reagents, such as poor ESI response.…”

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confidence: 99%

Basic Vapor Exposure for Tuning the Charge State Distribution of Proteins in Negative Electrospray Ionization: Elucidation of Mechanisms by Fluorescence Spectroscopy

Girod

Antoine

Dugourd

et al. 2012

J. Am. Soc. Mass Spectrom.

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Manipulation for simplifying or increasing the observed charge state distributions of proteins can be highly desirable in mass spectrometry experiments. In the present work, we implemented a vapor introduction technique to an Agilent Jet Stream ESI (Agilent Technologies, Santa Clara, CA, USA) source. An apparatus was designed to allow for the enrichment of the nitrogen sheath gas with basic vapors. An optical setup, using laser-induced fluorescence and a pH-chromic dye, permits the pH profiling of the droplets as they evaporate in the electrospray plume. Mechanisms of pH droplet modification and its effect on the protein charging phenomenon are elucidated. An important finding is that the enrichment with basic vapors of the nitrogen sheath gas, which surrounds the nebulizer spray, leads to an increase in the spray current. This is attributed to an increase in the electrical conductivity of water-amine enriched solvent at the tip exit. Here, the increased current results in a generation of additional electrolytically produced OH -ions and a corresponding increase in the pH at the tip exit. Along the electrospray plume, the pH of the droplets increases due to both droplet evaporation and exposure to basic vapors from the seeded sheath gas. The pH evolution in the ESI plume obtained using pure and basic seeded sheath gas was correlated with the evolution of the charge state distribution observed in mass spectra of proteins, in the negative ion mode. Taking advantage of the Agilent Jet Stream source geometry, similar protein charge state distributions and ion intensities obtained with basic initial solutions, can be obtained using native solution conditions by seeding the heated sheath gas with basic vapors.

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Shifts in protein charge state distributions with varying redox reagents in nanoelectrospray triple quadrupole mass spectrometry

Cited by 12 publications

References 55 publications

Ascorbic acid for homogenous redox buffering in electrospray ionization–mass spectrometry

Ascorbic acid for homogenous redox buffering in electrospray ionization–mass spectrometry

FT‐ICRMS distinguishes the mechanism of the charge state reduction for multiply charged protein cations admixed with redox reagents in ESI

Basic Vapor Exposure for Tuning the Charge State Distribution of Proteins in Negative Electrospray Ionization: Elucidation of Mechanisms by Fluorescence Spectroscopy

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