Glycomics and glycoproteomics: Approaches to address isomeric separation of glycans and glycopeptides

Reyes, Cristian D. Gutierrez; Jiang, Peilin; Donohoo, Kaitlyn; Atashi, Mojgan; Mechref, Yehia

doi:10.1002/jssc.202000878

Cited by 55 publications

(40 citation statements)

References 148 publications

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“…Glycopeptide separation prior to mass spectrometry detection, commonly performed using reversed-phase nano-scale LC, is an important component of most, if not all, glycoproteomics experiments. Current strategies for the isomeric separation of glycopeptides were recently reviewed [ 160 ].…”

Section: Innovations In Structure-focused Glycoproteomicsmentioning

confidence: 99%

Towards structure-focused glycoproteomics

Chernykh

Kawahara

Thaysen‐Andersen

2021

Biochemical Society Transactions

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Facilitated by advances in the separation sciences, mass spectrometry and informatics, glycoproteomics, the analysis of intact glycopeptides at scale, has recently matured enabling new insights into the complex glycoproteome. While diverse quantitative glycoproteomics strategies capable of mapping monosaccharide compositions of N- and O-linked glycans to discrete sites of proteins within complex biological mixtures with considerable sensitivity, quantitative accuracy and coverage have become available, developments supporting the advancement of structure-focused glycoproteomics, a recognised frontier in the field, have emerged. Technologies capable of providing site-specific information of the glycan fine structures in a glycoproteome-wide context are indeed necessary to address many pending questions in glycobiology. In this review, we firstly survey the latest glycoproteomics studies published in 2018–2020, their approaches and their findings, and then summarise important technological innovations in structure-focused glycoproteomics. Our review illustrates that while the O-glycoproteome remains comparably under-explored despite the emergence of new O-glycan-selective mucinases and other innovative tools aiding O-glycoproteome profiling, quantitative glycoproteomics is increasingly used to profile the N-glycoproteome to tackle diverse biological questions. Excitingly, new strategies compatible with structure-focused glycoproteomics including novel chemoenzymatic labelling, enrichment, separation, and mass spectrometry-based detection methods are rapidly emerging revealing glycan fine structural details including bisecting GlcNAcylation, core and antenna fucosylation, and sialyl-linkage information with protein site resolution. Glycoproteomics has clearly become a mainstay within the glycosciences that continues to reach a broader community. It transpires that structure-focused glycoproteomics holds a considerable potential to aid our understanding of systems glycobiology and unlock secrets of the glycoproteome in the immediate future.

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Section: Innovations In Structure-focused Glycoproteomicsmentioning

confidence: 99%

Towards structure-focused glycoproteomics

Chernykh

Kawahara

Thaysen‐Andersen

2021

Biochemical Society Transactions

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“…This was followed by permethylation that enhanced the glycan hydrophobicity, thereby increasing ionization efficiency and rendering them amenable to positive ionization. Permethylated glycans were also well retained in RP columns where their separation facilitated avoiding competitive ionization and simplified spectral interpretation [33]. This strategy was directly compared against the benchmark method (Cho et al [10]) using 50 μL of CSF and Filter-Aided N-Glycan Separation (FANGS) [35,36].…”

Section: Resultsmentioning

confidence: 99%

“…In the past, the comprehensive characterization of the CSF glycome has required several hundred microliters of fluid [22,32] and highly sensitive LC-MS analytical strategies. Additionally, glycan analysis using reverse-phase liquid chromatography and mass spectrometry (RPLC-MS) should be accompanied by a derivatization technique to overcome the poor retention of native glycans in common reverse phase (RP) columns, as well as their low ionization efficiency in positive mode [33].…”

Section: Introductionmentioning

confidence: 99%

N-Glycomics of Cerebrospinal Fluid: Method Comparison

2021

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Cerebrospinal fluid (CSF) contains valuable biological and neurological information. However, its glycomics analysis is hampered due to the low amount of protein in the biofluid, as has been demonstrated by other glycomics studies using a substantial amount of CSF. In this work, we investigated different N-glycan sample preparation approaches to develop a more sensitive method. These methods, one with an increased amount of buffer solution during the N-glycan release step with a lower amount of sample volume and the other with Filter-Aided N-Glycan Separation (FANGS), were compared with recent work to demonstrate their effectiveness. It was demonstrated that an increased amount of buffer solution showed higher intensity in comparison to the previously published method and FANGS. This suggested that digestion efficiency during the N-glycan release step was not in an optimal condition from the previously published method, and that there is a substantial loss of sample with FANGS when preparing N-glycans from CSF.

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“…Glycosidases, such as endo F3 and neuraminidase, are useful in studying specific glycan isomers, which have been shown to differ in pancreatic disease states. Isomeric characterization in glycoproteomics is a constantly evolving area of work and has been thoroughly reviewed recently (Gutierrez Reyes et al, 2021;Peng et al, 2021). Due to the heterogeneity of O-glycan core structures, a universal enzyme has not been discovered, though as shown by Malaker et al (2019), enzyme development for use in MS-based studies is a continued area of work.…”

Section: Future Perspectivesmentioning

confidence: 99%

Recent Advances in Mass Spectrometry-Based Glycomic and Glycoproteomic Studies of Pancreatic Diseases

Tabang

Ford

2021

Front. Chem.

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Modification of proteins by glycans plays a crucial role in mediating biological functions in both healthy and diseased states. Mass spectrometry (MS) has emerged as the most powerful tool for glycomic and glycoproteomic analyses advancing knowledge of many diseases. Such diseases include those of the pancreas which affect millions of people each year. In this review, recent advances in pancreatic disease research facilitated by MS-based glycomic and glycoproteomic studies will be examined with a focus on diabetes and pancreatic cancer. The last decade, and especially the last five years, has witnessed developments in both discovering new glycan or glycoprotein biomarkers and analyzing the links between glycans and disease pathology through MS-based studies. The strength of MS lies in the specificity and sensitivity of liquid chromatography-electrospray ionization MS for measuring a wide range of biomolecules from limited sample amounts from many sample types, greatly enhancing and accelerating the biomarker discovery process. Furthermore, imaging MS of glycans enabled by matrix-assisted laser desorption/ionization has proven useful in complementing histology and immunohistochemistry to monitor pancreatic disease progression. Advances in biological understanding and analytical techniques, as well as challenges and future directions for the field, will be discussed.

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Glycomics and glycoproteomics: Approaches to address isomeric separation of glycans and glycopeptides

Cited by 55 publications

References 148 publications

Towards structure-focused glycoproteomics

Towards structure-focused glycoproteomics

N-Glycomics of Cerebrospinal Fluid: Method Comparison

Recent Advances in Mass Spectrometry-Based Glycomic and Glycoproteomic Studies of Pancreatic Diseases

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