193‐nm photodissociation of singly and multiply charged peptide anions for acidic proteome characterization

Madsen, James A.; Kaoud, Tamer S.; Dalby, Kevin N.; Brodbelt, Jennifer S.

doi:10.1002/pmic.201000565

Cited by 75 publications

(147 citation statements)

References 14 publications

(20 reference statements)

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“…Recently, we investigated the utility of using 193-nm UVPD to sequence both singly and multiply charged peptide anions (20). This method generated rich and predictable fragmentation patterns, with a-/x-type products being the most prolific and abundant sequence ions, often yielding 100% sequence coverage.…”

Section: Resultsmentioning

confidence: 99%

“…All other reagents and solvents were obtained from Fisher Scientific (Fairlawn, NJ). An equimolar mixture of mitogen-activated protein kinases (MAPKs), including c-Jun N-terminal kinase 2, mitogen-activated protein kinase 1, and p38 mitogen-activated kinase 14, were prepared as described elsewhere (20). HeLa cells were obtained from American Type Culture Collection (CCL-2) (Manassas, VA) and were lysed, reduced, alkylated, and digested as reported previously (30).…”

Section: Methodsmentioning

confidence: 99%

“…In addition, the fragmentation patterns frequently yield many high-abundance neutral losses from product ions, which clutter the spectra (14 -17), and few sequence ions (14,18,19). Recently, we reported the use of 193-nm photons (ultraviolet photodissociation (UVPD)) for peptide anion activation, which was shown to yield rich and predictable fragmentation patterns with high sequence coverage on a fast liquid chromatographic timeline (20). This method showed promise for a range of peptide charge states (i.e.…”

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confidence: 99%

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High-throughput Database Search and Large-scale Negative Polarity Liquid Chromatography–Tandem Mass Spectrometry with Ultraviolet Photodissociation for Complex Proteomic Samples

Madsen

Robinson

et al. 2013

Molecular & Cellular Proteomics

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The use of ultraviolet photodissociation (UVPD) for the activation and dissociation of peptide anions is evaluated for broader coverage of the proteome. To facilitate interpretation and assignment of the resulting UVPD mass spectra of peptide anions, the MassMatrix database search algorithm was modified to allow automated analysis of negative polarity MS/MS spectra. The new UVPD algorithms were developed based on the MassMatrix database search engine by adding specific fragmentation pathways for UVPD. The new UVPD fragmentation pathways in MassMatrix were rigorously and statistically optimized using two large data sets with high mass accuracy and high mass resolution for both MS 1 and MS 2 data acquired on an Orbitrap mass spectrometer for complex Halobacterium and HeLa proteome samples. Negative mode UVPD led to the identification of 3663 and 2350 peptides for the Halo and HeLa tryptic digests, respectively, corresponding to 655 and 645 peptides that were unique when compared with electron transfer dissociation (ETD), higher energy collision-induced dissociation, and collision-induced dissociation results for the same digests analyzed in the positive mode. In sum, 805 and 619 proteins were identified via UVPD for the Halobacterium and HeLa samples, respectively, with 49 and 50 unique proteins identified in contrast to the more conventional MS/MS methods. The algorithm also features automated charge determination for low mass accuracy data, precursor filtering (including intact charge-reduced peaks), and the ability to combine both positive and negative MS/MS spectra into a single search, and it is freely open to the public. The accuracy and specificity of the MassMatrix UVPD search algorithm was also assessed for low resolution, low mass accuracy data on a linear ion trap. Analysis of a known mixture of three mitogen-activated kinases yielded similar sequence coverage percentages for UVPD of peptide anions versus conventional collisioninduced dissociation of peptide cations, and when these methods were combined into a single search, an increase of up to 13% sequence coverage was observed for the kinases. The ability to sequence peptide anions and cations in alternating scans in the same chromatographic run was also demonstrated. Because ETD has a significant bias toward identifying highly basic peptides, negative UVPD was used to improve the identification of the more acidic peptides in conjunction with positive ETD for the more basic species. In this case, tryptic peptides from the cytosolic section of HeLa cells were analyzed by polarity switching nanoLC-MS/MS utilizing ETD for cation sequencing and UVPD for anion sequencing. Relative to searching using ETD alone, positive/negative polarity switching significantly improved sequence coverages across identified proteins, resulting in a 33% increase in unique peptide identifications and more than twice the number of peptide spectral matches. Molecular & Cellular Proteomics 12: 10.1074/mcp.O113.028258, 2604-2614, 2013.The advent of new high-performance tandem mas...

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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High-throughput Database Search and Large-scale Negative Polarity Liquid Chromatography–Tandem Mass Spectrometry with Ultraviolet Photodissociation for Complex Proteomic Samples

Madsen

Robinson

et al. 2013

Molecular & Cellular Proteomics

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“…In particular, very recently the group of Brodbelt has implemented 193-nm ultraviolet photodissociation (UVPD) to sequence singly and multiply charged peptide anions. Upon dissociation by this method, a-/x-type, followed by d and w side-chain loss ions, were the most prolific and abundant sequence ions, often yielding 100% sequence coverage [26].…”

Section: Introductionmentioning

confidence: 99%

Formation and Fragmentation of Radical Peptide Anions: Insights from Vacuum Ultra Violet Spectroscopy

Brunet

Antoine

Dugourd

et al. 2011

J. Am. Soc. Mass Spectrom.

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We have studied the photodissociation of gas-phase deprotonated caerulein anions by vacuum ultraviolet (VUV) photons in the 4.5 to 20 eV range, as provided by the DESIRS beamline at the synchrotron radiation facility SOLEIL (France). Caerulein is a sulphated peptide with three aromatic residues and nine amide bonds. Electron loss is found to be the major relaxation channel at every photon energy. However, an increase in the fragmentation efficiency (neutral losses and peptide backbone cleavages) as a function of the energy is also observed. The oxidized ions, generated by electron photodetachment were further isolated and activated by collision (CID) in a MS 3 scheme. The branching ratios of the different fragments observed by CID as a function of the initial VUV photon energy are found to be independent of the initial photon energy. Thus, there is no memory effect of the initial excitation energy on the fragmentation channels of the oxidized species on the time scale of our tandem MS experiment. We also report photofragment yields as a function of photon energy for doubly deprotonated caerulein ions, for both closed-shell ([M -2H] 2-) non-radical ions and open-shell ([M -3H] 2-• ) radical ions. These latter ions are generated by electron photodetachment from [M -3H] 3-precursor ions. The detachment yield increases monotonically with the energy with the appearance of several absorption bands. Spectra for radical and non-radical ions are quite similar in terms of observed bands; however, the VUV fragmentation yield is enhanced by the presence of a radical in caerulein peptides.

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“…Direct infusion analysis on the LTQ XL was performed using an online nanoESI setup as previously described using a flow rate of 3 l/min at a concentration of 10 M in 49.5:49.5:1 MeOH:H 2 O:Acetic acid (44). An ESI voltage of 2 kV and a heated capillary temperature of 180°C were used for experiments on the LTQ XL.…”

Section: Methodsmentioning

confidence: 99%

Mapping Protein Surface Accessibility via an Electron Transfer Dissociation Selectively Cleavable Hydrazone Probe

Vasicek

O’Brien

Browning

et al. 2012

Molecular & Cellular Proteomics

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A protein's surface influences its role in protein-protein interactions and protein-ligand binding. Mass spectrometry can be used to give low resolution structural information about protein surfaces and conformations when used in combination with derivatization methods that target surface accessible amino acid residues. However, pinpointing the resulting modified peptides upon enzymatic digestion of the surface-modified protein is challenging because of the complexity of the peptide mixture and low abundance of modified peptides. Here a novel hydrazone reagent (NN) is presented that allows facile identification of all modified surface residues through a preferential cleavage upon activation by electron transfer dissociation coupled with a collision activation scan to pinpoint the modified residue in the peptide sequence.

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193‐nm photodissociation of singly and multiply charged peptide anions for acidic proteome characterization

Cited by 75 publications

References 14 publications

High-throughput Database Search and Large-scale Negative Polarity Liquid Chromatography–Tandem Mass Spectrometry with Ultraviolet Photodissociation for Complex Proteomic Samples

High-throughput Database Search and Large-scale Negative Polarity Liquid Chromatography–Tandem Mass Spectrometry with Ultraviolet Photodissociation for Complex Proteomic Samples

Formation and Fragmentation of Radical Peptide Anions: Insights from Vacuum Ultra Violet Spectroscopy

Mapping Protein Surface Accessibility via an Electron Transfer Dissociation Selectively Cleavable Hydrazone Probe

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