Solution Additives that Desalt Protein Ions in Native Mass Spectrometry

Flick, Tawnya G.; Cassou, Catherine A.; Chang, Terrence M.; Williams, Evan R.

doi:10.1021/ac301629s

Cited by 60 publications

(69 citation statements)

References 45 publications

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“…Ammonium salts have also been used in tissue analysis providing a 5× increase in the total ion current and improvement of signal-to-noise using negative ion MALDI enhancing imaging of glycerophospholipids, sulfatides, and gangliosides [25]. High concentrations of ammonium salts have also been used in ESI for the analysis of protein complexes, preventing protein unfolding and maintaining the complexes intact, allowing so called native MS [26][27][28]. Certain ammonium salts also remove cation adduct formation in ESI [28,29].…”

Section: Introductionmentioning

confidence: 99%

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Improving the Sensitivity of Matrix-Assisted Ionization (MAI) Mass Spectrometry Using Ammonium Salts

Chubatyi

McEwen

2015

J. Am. Soc. Mass Spectrom.

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Abstract. In matrix-assisted ionization (MAI), analyte incorporated in a small molecule matrix is introduced into an aperture linking atmospheric pressure with the vacuum of a mass spectrometer. Gas-phase analyte ions are spontaneously produced without use of a laser or high voltage. Here we investigate analyte and background ion abundances upon addition of ammonium salts to various MAI matrix/analyte solutions. Regardless of the ammonium salt or matrix used, chemical background ions are suppressed and/or analyte ion abundance improved for basic small molecules, peptides, and proteins. Background ion abundances increase with increasing inlet temperature, but are suppressed with addition of any of a variety of ammonium salts without much effect on the total ion abundances of the analyte ions. However, at lower inlet temperature using the matrices 2-bromo-2-nitropropane-1,3-diol, 1,2-dicyanobenzene, and 3-nitrobenzonitrile (3-NBN), analyte ion abundance increases and any chemical background decreases upon addition of ammonium salt. The improvement in sensitivity using 3-NBN with ammonium salt allows full acquisition mass spectra consuming as little as 1 fmol of ubiquitin. More complete peptide coverage for 100 fmol of a BSA tryptic digest, and increased sensitivity of drugs spiked in urine and saliva were observed after ammonium salt addition to the 3-NBN matrix.

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Section: Introductionmentioning

confidence: 99%

“…High concentrations of ammonium salts have also been used in ESI for the analysis of protein complexes, preventing protein unfolding and maintaining the complexes intact, allowing so called native MS [26][27][28]. Certain ammonium salts also remove cation adduct formation in ESI [28,29].…”

Section: Introductionmentioning

confidence: 99%

Improving the Sensitivity of Matrix-Assisted Ionization (MAI) Mass Spectrometry Using Ammonium Salts

Chubatyi

McEwen

2015

J. Am. Soc. Mass Spectrom.

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“…23 McLuckey and coworkers found that the extent of sodium ion adduction on a protein ion formed by ESI is related to the protein pI, solution pH, and charge state. 18, 26 There is more sodium ion adduction to low charge states, 18, 21, 25–27 but there is more adduction of trivalent metal ions to high charge states. 28 …”

Section: Introductionmentioning

confidence: 99%

“…For example, 25 mM ammonium bromide added to an aqueous 1 mM NaCl solution containing ubiquitin decreases the average number of sodium ions adducted to the most abundant charge state of the protein from 6.0 to 0.4 and increases the S/N of this ion by a factor of 66. 25, 27 The ability of different salts to desalt proteins in the ESI droplet is related to the proton affinities of the anions, where anions with low proton affinities lead to less sodium adduction. 27 However, anions with low proton affinities also tend to adduct to the protein as an acid molecule and form ion clusters, 27 which can decrease the protein ion signal.…”

Section: Introductionmentioning

confidence: 99%

Desalting protein ions in native mass spectrometry using supercharging reagents

Cassou

Williams

2014

Analyst

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Effects of the supercharging reagents m-NBA and sulfolane on sodium ion adduction to protein ions formed using native mass spectrometry were investigated. There is extensive sodium adduction on protein ions formed by electrospray ionization from aqueous solutions containing millimolar concentrations of NaCl, which can lower sensitivity by distributing the signal of a given charge state over multiple adducted ions and can reduce mass measuring accuracy for large proteins and non-covalent complexes for which individual adducts cannot be resolved. The average number of sodium ions adducted to the most abundant ion formed from ten small (8.6–29 kDa) proteins for which adducts can be resolved is reduced by 58% or 80% on average, respectively, when 1.5% m-NBA or 2.5% sulfolane are added to aqueous solutions containing sodium compared to without the supercharging reagent. Sulfolane is more effective than m-NBA at reducing sodium ion adduction and at preserving non-covalent protein-ligand and protein-protein interactions. Desalting with 2.5% sulfolane enables detection of several glycosylated forms of 79.7 kDa holo-transferrin and NADH bound to the 146 kDa homotetramer LDH, which are otherwise unresolved due to peak broadening from extensive sodium adduction. Although sulfolane is more effective than m-NBA at protein ion desalting, m-NBA reduces salt clusters at high m/z and can increase the signal-to-noise ratios of protein ions by reducing chemical noise. Desalting is likely a result of these supercharging reagents binding sodium ions in solution, thereby reducing the sodium available to adduct to protein ions.

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“…Several strategies have been developed that enable the presence of mM concentrations of non-volatile salts in nano-ESI analyses. These include adding supercharging reagents [65], additives [66][67][68], or high ammonium acetate concentrations to prevent salt-induced ion suppression/adduction [60,69,70]. More recently it has been shown that using submicrometer nano-ESI capillaries (<0.5 mm compared with conventional emitter sizes of 1-10 lm) enable the detection of well-resolved charge state distributions for native proteins and protein complexes analyzed from highsalt biochemical buffers (e.g.…”

Section: Introductionmentioning

confidence: 99%

Mass spectrometry-enabled structural biology of membrane proteins

Calabrese

Radford

2018

Methods

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a b s t r a c tThe last $25 years has seen mass spectrometry (MS) emerge as an integral method in the structural biology toolkit. In particular, MS has enabled the structural characterization of proteins and protein assemblies that have been intractable by other methods, especially those that are large, heterogeneous or transient, providing experimental evidence for their structural organization in support of, and in advance of, high resolution methods. The most recent frontier conquered in the field of MS-based structural biology has been the application of established methods for studying water soluble proteins to the more challenging targets of integral membrane proteins. The power of MS in obtaining structural information has been enabled by advances in instrumentation and the development of hyphenated mass spectrometry-based methods, such as ion mobility spectrometry-MS, chemical crosslinking-MS and other chemical labelling/footprinting-MS methods. In this review we detail the insights garnered into the structural biology of membrane proteins by applying such techniques. Application and refinement of these methods has yielded unprecedented insights in many areas, including membrane protein conformation, dynamics, lipid/ligand binding, and conformational perturbations due to ligand binding, which can be challenging to study using other methods.

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Solution Additives that Desalt Protein Ions in Native Mass Spectrometry

Cited by 60 publications

References 45 publications

Improving the Sensitivity of Matrix-Assisted Ionization (MAI) Mass Spectrometry Using Ammonium Salts

Improving the Sensitivity of Matrix-Assisted Ionization (MAI) Mass Spectrometry Using Ammonium Salts

Desalting protein ions in native mass spectrometry using supercharging reagents

Mass spectrometry-enabled structural biology of membrane proteins

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