Site-Specific Glycosylation Quantitation of 50 Serum Glycoproteins Enhanced by Predictive Glycopeptidomics for Improved Disease Biomarker Discovery

Li, Qiongyu; Kailemia, Muchena J.; Merleev, Alexander A.; Xu, Gege; Serie, Daniel J.; Danan, Lieza M.; Haj, Fawaz G.; Maverakis, Emanual; Lebrilla, Carlito B.

doi:10.1021/acs.analchem.9b00776

Cited by 47 publications

(52 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Glycoproteins are well‐known to be associated with diseases (especially cancers) and were used for discovering biomarkers in lung cancer plasma . We worked on the plasma derived from lung cancer patients with the glycoproteomics approach, classified the identified glycoproteins, and comparatively analyzed them to elucidate the relationships of the identified proteins.…”

Section: Resultsmentioning

confidence: 99%

Glycoproteomics Method to Discover Reliable Biomarkers from Human Plasma of Lung Cancer Patients for MS‐based Clinical Studies

Lee

Shin

Jeong

et al. 2019

Bulletin Korean Chem Soc

View full text Add to dashboard Cite

In clinical and diagnostic proteomics, it is important to discover significant biomarkers from biosamples. Thus, a reliable proteomics methodology is required for the development and standardization of an MSbased protein identification and quantification method in biosamples. In particular, plasma is one of the most complex fluids in the human body and is commonly used in hospitals to diagnose disease. The purpose of this study was to develop a reliable and reproducible proteomics method using lung cancer plasma for biomarker discovery and diagnostic kits for screening. Glycoproteins are well-known to be associated with diseases (especially cancers) so in this study glycoproteomics was used for discovering biomarkers from lung cancer plasma. Thirty-five lung cancer plasma were pooled, and glycoproteins were separated using lectin affinity chromatography (LAC). After affinity selection, trypsin-digestion, and deglycosylation with PNGase F, the resulting deglycosylated peptides were analyzed with nLC-MS/MS runs twice. The corresponding parent proteins were identified separately through two database search engines and then analyzed individually. The identified proteins from each set were compared, combined, and then categorized for analysis. The combined total identified number of proteins were about 50% increased than in a single run and some proteins seemed to be more reproducible and reliable biomarker candidates because they were always identified in every run. From this research, method-optimized proteomics can be applied to biomarker discovery for diagnosis and prognosis of disease such as cancer for better MS-based clinical studies. The identified plasma proteins from this research will also be lung cancer biomarker candidates and can be utilized for the development of in vitro lung cancer diagnostic kits.

show abstract

Section: Resultsmentioning

confidence: 99%

Glycoproteomics Method to Discover Reliable Biomarkers from Human Plasma of Lung Cancer Patients for MS‐based Clinical Studies

Lee

Shin

Jeong

et al. 2019

Bulletin Korean Chem Soc

View full text Add to dashboard Cite

show abstract

“…The neat enzymatically-prepared samples containing both peptides and glycopeptides were then directly analyzed without further hands-on sample cleanup or dilution using an Agilent 1290 infinity liquid chromatography (LC) system coupled to an Agilent 6490 triple quadrupole (QqQ) mass spectrometer (Agilent Technologies, Santa Clara, CA), as previously described 23,35,36 . Briefly, an Agilent Eclipse plus C18 (RRHD 1.8 µm, 2.1 × 100 mm) coupled with an Agilent Eclipse plus C18 pre-column (RRHD 1.8 µm, 2.1 × 5 mm) was used for UPLC separation.…”

Section: Uplc-esi-qqq-ms Analysismentioning

confidence: 99%

“…The MRM MS method used for this study requires predetermined knowledge of the peptide or glycopeptide's LC retention time and its collision induced dissociation (CID) behavior, which we have previously determined for all the non-glycosylated peptides and glycopeptides used in this study ( Fig. S8 and Table S1) 17,35,36 . The specific method used herein has been highly validated and the monitored transitions have been described in detail 16,17,36 .…”

Section: Uplc-esi-qqq-ms Analysismentioning

confidence: 99%

See 1 more Smart Citation

A site-specific map of the human plasma glycome and its age and gender-associated alterations

Merleev¹,

Park²,

Xie³

et al. 2020

Sci Rep

Self Cite

View full text Add to dashboard Cite

Alterations in the human glycome have been associated with cancer and autoimmunity. Thus, constructing a site-specific map of the human glycome for biomarker research and discovery has been a highly sought-after objective. However, due to analytical barriers, comprehensive site-specific glycoprofiling is difficult to perform. To develop a platform to detect easily quantifiable, site-specific, disease-associated glycan alterations for clinical applications, we have adapted the multiple reaction monitoring mass spectrometry method for use in glycan biomarker research. The adaptations allow for highly precise site-specific glycan monitoring with minimum sample prep. Using this technique, we successfully mapped out the relative abundances of the most common 159 glycopeptides in the plasma of 97 healthy volunteers. This plasma glycome map revealed 796 significant (FDR < 0.05) site-specific inter-protein and intra-protein glycan associations, of which the vast majority were previously unknown. Since age and gender are relevant covariants in biomarker research, these variables were also characterized. 13 glycopeptides were found to be associated with gender and 41 to be associated with age. Using just five age-associated glycopeptides, a highly accurate age prediction model was constructed and validated (r2 = 0.62 ± 0.12). The human plasma site-specific glycan map described herein has utility in applications ranging from glycan biomarker research and discovery to the development of novel glycan-altering interventions.

show abstract

“…[33][34][35][36][37][38][39][40][41][42][43][44][45] Even so, HCD remains widely used in N-glycoproteomics. [46][47][48][49][50][51][52][53][54][55][56][57] Tryptic Nglycopeptides tend to harbor only one potential glycosite, as defined by its sequon N-X-S/T, where X represents any amino acid other than proline, which limits dependence on peptide fragments that retain intact glycans. HCD of N-glycopeptides also often generates b/y-type fragments that retain a N-acetylglucosamine (GlcNAc) moiety that provide clues to glycosite localization.…”

Section: Introductionmentioning

confidence: 99%

Optimal Dissociation Methods Differ for N- and O-glycopeptides

Riley

Malaker²,

Drießen³

et al. 2020

Preprint

View full text Add to dashboard Cite

<p><a>Site-specific characterization of glycosylation requires intact glycopeptide analysis, and recent efforts have focused on how to best interrogate glycopeptides using tandem mass spectrometry (MS/MS). Beam-type collisional activation, i.e., higher-energy collisional dissociation (HCD), has been a valuable approach, but stepped collision energy HCD (sceHCD) and electron transfer dissociation with HCD supplemental activation (EThcD) have emerged as potentially more suitable alternatives. Both sceHCD and EThcD have been used with success in large-scale glycoproteomic experiments, but they each incur some degree of compromise. Most progress has occurred in the area N-glycoproteomics. There is growing interest in extending this progress to O-glycoproteomics, which necessitates comparisons of method performance for the two classes of glycopeptides. Here, we systematically explore the advantages and disadvantages of conventional HCD, sceHCD, ETD, and EThcD for intact glycopeptide analysis and determine their suitability for both N- and O-glycoproteomic applications. For N-glycopeptides, HCD and sceHCD generate similar numbers of identifications, although sceHCD generally provides higher quality spectra. Both significantly outperform EThcD methods, indicating that ETD-based methods are not required for routine N-glycoproteomics. Conversely, ETD-based methods, especially EThcD, are indispensable for site-specific analyses of O-glycopeptides. Our data show that O-glycopeptides cannot be robustly characterized with HCD-centric methods that are sufficient for N-glycopeptides, and glycoproteomic methods aiming to characterize O-glycopeptides must be constructed accordingly.</a></p>

show abstract

Site-Specific Glycosylation Quantitation of 50 Serum Glycoproteins Enhanced by Predictive Glycopeptidomics for Improved Disease Biomarker Discovery

Cited by 47 publications

References 34 publications

Glycoproteomics Method to Discover Reliable Biomarkers from Human Plasma of Lung Cancer Patients for MS‐based Clinical Studies

Glycoproteomics Method to Discover Reliable Biomarkers from Human Plasma of Lung Cancer Patients for MS‐based Clinical Studies

A site-specific map of the human plasma glycome and its age and gender-associated alterations

Optimal Dissociation Methods Differ for N- and O-glycopeptides

Contact Info

Product

Resources

About