High-resolution electrospray mass spectra of large molecules

Henry, Kent D.; Quinn, John; McLafferty, Fred W.

doi:10.1021/ja00014a043

Cited by 110 publications

(74 citation statements)

References 0 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Designated preliminary experiments were run on a similar instrument with a 2.8-T magnet as described (5,6). The ions were allowed to react for different time periods (separate runs) with 2H20 before excitation and detection.…”

Section: Methodsmentioning

confidence: 99%

“…Protein folding is probably the least well understood step in the sequence of transformations relating genetic information with its expression by protein function (1). Dramatic new ionization methods for mass spectrometry (MS) have made possible the formation of protein ions in the gas phase to measure molecular weight and primary sequence information (2-4), even on fmol samples (5,6). Recent studies indicate that protein conformations in solution can affect the resulting charge distribution of the gaseous multiply charged ions formed by electrospray ionization (ESI) (7)(8)(9) and that even noncovalent complexes can survive ESI to form gaseous multiply charged ions (10)(11)(12)(13)(14)(15).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Coexisting stable conformations of gaseous protein ions.

Suckau

Shi

Beu

et al. 1993

Proc. Natl. Acad. Sci. U.S.A.

345

308

View full text Add to dashboard Cite

For further insight into the role of solvent in protein conformer stabilization, the structural and dynamic properties of protein ions in vacuo have been probed by hydrogen-deuterium exchange in a Fourier-transform mass spectrometer. Multiply charged ions generated by electrospray ionization of five proteins show exchange reactions with 2H20 at 10-7 torr (1 torr = 133.3 Pa) exhibiting pseudo-first-order kinetics. Gas-phase compactness of the S-S cross-linked RNase A relative to denatured S-derivatized RNase A is indicated by exchange of 35 and 135 hydrogen atoms, respectively. For pure cytochrome c ions, the existence of at least three distinct gaseous conformers is indicated by the substantially different values-52, 113, and 74 -ofreactive H atoms; the observation of these same values for ions of a number-2, 7, and 5, respectively-of different charge states indicates conformational insensitivity to coulombic forces. For each of these conformers, the compactness in vacuo indicated by these values corresponds directly to that of a known conformer structure in the solution from which the conformer ions are produced by electrospray. S-derivatized RNase A ions also exist as at least two gaseous conformers exchanging 50-140 H atoms. Gaseous conformer ions are isomerically stable for hours; removal of solvent greatly increases conformational rigidity. More specific ion-molecule reactions could provide further details of conformer structures.The relationship between the dynamic structure ofproteins in solution and their biological activity has been of longstanding research interest. Protein folding is probably the least well understood step in the sequence of transformations relating genetic information with its expression by protein function (1). Dramatic new ionization methods for mass spectrometry (MS) have made possible the formation of protein ions in the gas phase to measure molecular weight and primary sequence information (2-4), even on fmol samples (5, 6). Recent studies indicate that protein conformations in solution can affect the resulting charge distribution of the gaseous multiply charged ions formed by electrospray ionization (ESI) (7-9) and that even noncovalent complexes can survive ESI to form gaseous multiply charged ions (10)(11)(12)(13)(14)(15).Critical information concerning solvent effects on the conformation and dynamic properties of proteins has come from NMR (16) and from isotope-exchange experiments with 2H20 (17), including those before and during ESI/MS (18,19). With an activation energy of 17-20 kcal/mol (1 cal = 4.184 J) (17), the H/2H exchange rate depends on the pH (17), electrostatic effects (20), proximity of the solvent-accessible surface (21), and conformational flexibility with hydrogen bond cleavage and formation during local unfolding and folding (22). Studies ofgaseous proteins should help delineate the role of solvent in stabilizing protein conformations, but such previous studies have been mainly theoretical (23) because of the lack of experimental approaches. We repor...

show abstract

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

Coexisting stable conformations of gaseous protein ions.

Suckau

Shi

Beu

et al. 1993

Proc. Natl. Acad. Sci. U.S.A.

345

308

View full text Add to dashboard Cite

show abstract

“…A pulse of N2 gas is admitted to the ion cell to cool the ions translationally before measurement; other experimental details were as given (21)(22)(23).…”

Section: Methodsmentioning

confidence: 99%

“…This also benefits tandem mass spectrometry; dissociation of peptide ions as large as 8.5 kDa yields fragment masses consistent (<0. (21)(22)(23)(24)(25)(26) can be used to measure simultaneously all such ions as large as 17 kDa with 50,000-80,000 resolving power and, by using an internal standard, with <0.1 Da (<6 ppm) mass errors. As reported here, this enhanced high-resolution capability is also valuable for characterizing impurities (6-8) adducts (9-11) and, by tandem mass spectrometry (MS/MS) (3,7,8,(12)(13)(14)(15), amino acid sequences.…”

Section: Introductionmentioning

confidence: 99%

High-resolution tandem mass spectrometry of large biomolecules.

Loo

Quinn

Ryu

et al. 1992

Proc. Natl. Acad. Sci. U.S.A.

121

View full text Add to dashboard Cite

Unit-resolution mass spectra have been obtained for peptides as large as 17 kDa, providing information on impurities and adduct ions, as well as accurate molecular weight values. Electrospray ionization produces many multiply-charged species of the same mass; isotopic peak resolution provides direct charge state assignment from the unit mass spacing of the isotopes. This is of special value when the spectrum also has many masses, such as from precursor ion dissociation or impurities. Mass measuring errors not only are concomitantly lower (<0.1 Da) than when the isotopic peaks are unresolved but also are independent of variations in '3C/12C natural isotopic abundances. Also, larger errors are avoided that occur when the measured peak envelope includes impurity or adduct ions. This also benefits tandem mass spectrometry; dissociation of peptide ions as large as 8.5 kDa yields fragment masses consistent (<0. (21)(22)(23)(24)(25)(26) can be used to measure simultaneously all such ions as large as 17 kDa with 50,000-80,000 resolving power and, by using an internal standard, with <0.1 Da (<6 ppm) mass errors. As reported here, this enhanced high-resolution capability is also valuable for characterizing impurities (6-8) adducts (9-11) and, by tandem mass spectrometry (MS/MS) (3,7,8,(12)(13)(14)(15), amino acid sequences.EXPERIMENTAL PROCEDURES Radio frequency-only quadrupole lenses and five stages of differential pumping are used to transport the ions electrosprayed at -103 torr to the FTMS ion-measurement cell at <10-8 torr in a 2.8-T magnet. A pulse of N2 gas is admitted to the ion cell to cool the ions translationally before measurement; other experimental details were as given (21-23).RESULTS AND DISCUSSION and the measured value (weighted average of isotopic peak values; external standard: gramicidin S, Mr = 1141) is 12,352.20 (12,358.22 for the most abundant isotopic peak).Note that a 5-per-mil shiftt in the 13C/'2C ratio would change the isotopically averaged molecular weight (centroid of the unresolved isotopic peaks) (1-18) by 0.3 Da but would only affect the abundances of the isotopic peaks, not their masses. These experimental procedures have not produced, to date, high-resolution ESI spectra of molecules as large as albumin (66 kDa), although its spectrum (Fig. 2) was improved (22).By using a quadrupole mass spectrometer (6, 7), the ESI mass spectra of bovine ubiquitin, mMr 8559.62, showed peaks of measured masses 114.7 Da (6) and 115.1 Da (7) below those of the (M + nH)n+ ubiquitin peaks; § this was postulated (7) to result from an impurity missing the two carboxyl-terminal glycines, which would lower the mass by 114.04 Da. The ESI/FTMS mass spectrum of the same sample (7) (Fig. 3) shows an mMr of 8559.45 and peaks presumably from the same impurity at a mMr of 8445.43 (Am = 114.02). Additional anomalous peaks not resolved in the quadrupole spectra (6,7) give an average mMr of 8623.39 (Am = 63.94); replacing a valine by a tyrosine would increase the molecular weight by 63.99. As noted (23), ident...

show abstract

“…Due to the high mass accuracy and resolving power of an FTMS, MALDI-FTMS [5][6][7] and ESI-FTMS [8,9] are becoming the instruments of choice for proteomics [10], and experiments on proteins and large fragments of proteins, so called "top-down" [11][12][13][14] mass spectrometry. These experiments tend to slow down due to the lack of sophisticated methods for automatic spectrum analysis.…”

Section: T He Wide Employment Of Fourier Transform Massmentioning

confidence: 99%

Algorithms for automatic interpretation of high resolution mass spectra

Kaur

O’Connor

2006

J. Am. Soc. Mass Spectrom.

View full text Add to dashboard Cite

Automated interpretation of high-resolution mass spectra in a reliable and efficient manner represents a highly challenging computational problem. This work aims at developing methods for reducing a high-resolution mass spectrum into its monoisotopic peak list, and automatically assigning observed masses to known fragment ion masses if the protein sequence is available. The methods are compiled into a suite of data reduction algorithms which is called MasSPIKE (Mass Spectrum Interpretation and Kernel Extraction). MasSPIKE includes modules for modeling noise across the spectrum, isotopic cluster identification, charge state determination, separation of overlapping isotopic distributions, picking isotopic peaks, aligning experimental and theoretical isotopic distributions for estimating a monoisotopic peak's location, generating the monoisotopic mass list, and assigning the observed monoisotopic masses to possible protein fragments. The method is tested against a complex top-down spectrum of bovine carbonic anhydrase. Results of each of the individual modules are compared with previously published work. (J Am Soc Mass Spectrom 2006, 17, 459 -468)

show abstract

High-resolution electrospray mass spectra of large molecules

Cited by 110 publications

References 0 publications

Coexisting stable conformations of gaseous protein ions.

Coexisting stable conformations of gaseous protein ions.

High-resolution tandem mass spectrometry of large biomolecules.

Algorithms for automatic interpretation of high resolution mass spectra

Contact Info

Product

Resources

About