GlycoPep Detector: A Tool for Assigning Mass Spectrometry Data of N-Linked Glycopeptides on the Basis of Their Electron Transfer Dissociation Spectra

Zhu, Zhongkui; Hua, David; Clark, Daniel F.; Go, Eden P.; Desaire, Heather

doi:10.1021/ac400287n

Cited by 49 publications

(51 citation statements)

References 42 publications

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“…An example is given in Figure 6, where asialofetuin glycopeptides of varying charge states were fragmented by ETD. The c and z ions that result from peptide backbone cleavages are much more pronounced when the glycopeptide precursor is in the 6+ charge state, whereas no fragment ions except the charge-reduced precursors are found for the same glycopeptide in the 4+ charge state (97). To enhance the fragmentation efficiency in ETD, mnitrobenzyl alcohol, a supercharging reagent, was added to the LC eluate after column separation to increase the average charge state of glycopeptides (98).…”

Section: Electron-based Fragmentationmentioning

confidence: 99%

“…In view of the great complementarity possible with ETD, a few programs have included functionality to analyze the ETD data of glycopeptides (97,(115)(116)(117)(118). This functionality is particularly useful in O-linked glycopeptide analysis, because no consensus sequence exists to predict the potential O-linked glycosylation site, which is also hard to determine by CID data (119-121).…”

Section: Automated Glycoproteomicsmentioning

confidence: 99%

“…The software allows merging of CID and ETD search results, and users can compare different modification site assignments for site localization. To automate the glycopeptide identification workflow by ETD, Zhu et al (97,117) studied the fragmentation patterns of N-and O-linked glycopeptides from model glycoproteins and developed novel algorithms to consider multiple ion series (c, z, and y ions) of putative glycopeptide candidates separately. By implementing the algorithms into standalone software programs that handle glycopeptide ETD data, the correct glycopeptide compositions and site assignments were made with high throughput and high accuracy.…”

Section: Automated Glycoproteomicsmentioning

confidence: 99%

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Carbohydrates on Proteins: Site-Specific Glycosylation Analysis by Mass Spectrometry

Zhu

Desaire²

2015

Annual Rev. Anal. Chem.

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Glycosylation on proteins adds complexity and versatility to these biologically vital macromolecules. To unveil the structure-function relationship of glycoproteins, glycopeptide-centric analysis using mass spectrometry (MS) has become a method of choice because the glycan is preserved on the glycosylation site and site-specific glycosylation profiles of proteins can be readily determined. However, glycopeptide analysis is still challenging given that glycopeptides are usually low in abundance and relatively difficult to detect and the resulting data require expertise to analyze. Viewing the urgent need to address these challenges, emerging methods and techniques are being developed with the goal of analyzing glycopeptides in a sensitive, comprehensive, and high-throughput manner. In this review, we discuss recent advances in glycoprotein and glycopeptide analysis, with topics covering sample preparation, analytical separation, MS and tandem MS techniques, as well as data interpretation and automation.

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Section: Electron-based Fragmentationmentioning

confidence: 99%

Section: Automated Glycoproteomicsmentioning

confidence: 99%

Section: Automated Glycoproteomicsmentioning

confidence: 99%

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Carbohydrates on Proteins: Site-Specific Glycosylation Analysis by Mass Spectrometry

Zhu

Desaire²

2015

Annual Rev. Anal. Chem.

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“…One software tool to aid in the interpretation of glycopeptide fragmentation spectra is GlycoMiner, an algorithm intended to approach MS/MS interpretation in a fashion similar to a human expert [137]. Desaire and coworkers have extended their work on GlycoPep DB, resulting in the more recently reported GlycoPep Grader, GlycoPep Detector and GlycoPep Scorer tools [138][139][140]. Collectively, these utilities provide support for interpretation of CID and ETD data for both N-linked and O-linked glycopeptide analytes.…”

Section: Tools For Interpretation Of Ms/ms Datamentioning

confidence: 99%

Engaging Challenges in Glycoproteomics: Recent Advances in MS-Based Glycopeptide Analysis

2015

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The proteomic analysis of glycosylation is uniquely challenging. The numerous and varied biological roles of protein-linked glycans have fueled a tremendous demand for technologies that enable rapid, in-depth structural examination of glycosylated proteins in complex biological systems. In turn, this demand has driven many innovations in wide ranging fields of bioanalytical science. This review will summarize key developments in glycoprotein separation and enrichment, glycoprotein proteolysis strategies, glycopeptide separation and enrichment, the role of mass measurement accuracy in glycopeptide detection, glycopeptide ion dissociation methods for MS/MS, and informatic tools for glycoproteomic analysis. In aggregate, this selection of topics serves to encapsulate the present status of MS-based analytical technologies for engaging the challenges of glycoproteomic analysis.

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“…At the commencement of this work most available software relied on pre-defined glycan databases (255,261,263,(271)(272)(273), in a manner analogous to proteomic software. Software that did not require prior knowledge of glycan databases [reviewed in (244) In this work, OxoExtract was often used several times to search a single mzML file.…”

Section: Discussionmentioning

confidence: 99%

Structural characterisation of glycosylation of paramyxovirus attachment and fusion proteins by mass spectrometry

Pegg¹

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The family Paramyxoviridae (paramyxovirus) contains several significant human and animal pathogens.Represented within this family are human respiratory syncytial virus (hRSV) and Newcastle disease virus (NDV). The former contributes significantly to severe respiratory tract disease in infants, children and immunocompromised individuals. At present, highly efficacious therapeutics or safe and effective vaccines are not available for hRSV. NDV is the causative agent of Newcastle disease, afflicting a wide range of avian species. The desire to study NDV is due not only to the significant economic impact it has on the poultry industry worldwide, but also its potential use as an oncolytic agent and vaccine vector in humans and animals. Additionally, findings on NDV may be translated to closely related viruses that cause disease in humans, such as parainfluenza viruses.As outlined in Chapter 1 of this thesis, the infectious processes of all members of this family are driven by two major membrane glycoproteins whose ectodomains project from the viral envelope: the fusion (F) and attachment glycoproteins. The F glycoprotein is responsible for viral entry by means of fusion with host cell membranes while the variable attachment glycoproteins, haemagglutinin, haemagglutininneuraminidase (HN) and major surface glycoprotein (G), are involved in viral attachment to host cells.Previous studies have shown that altering the glycosylation profile of these proteins can modulate the ability of the virus to infect host cells and stimulate the host immune system. As yet, site-specific glycan heterogeneity of hRSV and NDV surface glycoproteins has not been defined at a chemical level. As described in Chapter 2, reverse-phase liquid chromatography tandem mass spectrometry (MS) strategies were implemented to characterise intact glycopeptides from the attachment and F glycoproteins of hRSV and NDV. Site-occupancy and monosaccharide compositions at a given site were determined using collision-induced dissociation, higher-energy collision dissociation, electron-transfer dissociation and electron-transfer dissociation combined with collision dissociation. As described in Chapter 3, a spectral processing program was developed called OxoExtract to aid identification of intact glycopeptides that were fragmented with higher-energy collision dissociation.The F and HN proteins of NDV were derived from virions propagated in embryonic eggs. Analyses of HN in Chapter 4 revealed high mannose N-linked glycans and complex or hybrid N-linked glycans that were variably fucosylated, sialylated and sulfated or phosphorylated. In total 63, 58, and 37 glycans were identified at sites N341, N433 and N481, respectively. In addition, a previously undocumented Olinked glycosylation site was identified in the stalk domain of the protein. Observed glycans from NDV F described in Chapter 5 were mainly high mannose, containing variations of five to nine mannose ii residues across four sites, N85, N191, N366 and N471. There was also evidence of fucosylated c...

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GlycoPep Detector: A Tool for Assigning Mass Spectrometry Data of N-Linked Glycopeptides on the Basis of Their Electron Transfer Dissociation Spectra

Cited by 49 publications

References 42 publications

Carbohydrates on Proteins: Site-Specific Glycosylation Analysis by Mass Spectrometry

Carbohydrates on Proteins: Site-Specific Glycosylation Analysis by Mass Spectrometry

Engaging Challenges in Glycoproteomics: Recent Advances in MS-Based Glycopeptide Analysis

Structural characterisation of glycosylation of paramyxovirus attachment and fusion proteins by mass spectrometry

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