Assessment of lectin and HILIC based enrichment protocols for characterization of serum glycoproteins by mass spectrometry

Calvano, Cosima Damiana; Zambonin, Carlo; Jensen, Ole N.

doi:10.1016/j.jprot.2008.06.013

Cited by 122 publications

(114 citation statements)

References 49 publications

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“…Glycosylation is usually heterogeneous and a single peptide peak splits into multiple signals, accompanied by reduced signal intensity and altered retention on separation systems when compared with its unglycosylated counterpart (35). Furthermore, glycopeptide signals can be suppressed in the presence of peptides as a result of competitive ionization (45)(46)(47). Therefore, reduction of the sample complexity by a selective enrichment of glycopeptides is often beneficial.…”

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confidence: 99%

A Microarray-Matrix-assisted Laser Desorption/Ionization-Mass Spectrometry Approach for Site-specific Protein N-glycosylation Analysis, as Demonstrated for Human Serum Immunoglobulin M (IgM)*

Pabst

Kuster

Wahl³

et al. 2015

Molecular & Cellular Proteomics

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We demonstrate a new approach for the site-specific identification and characterization of protein N-glycosylation. It is based on a nano-liquid chromatography microarray-matrix assisted laser desorption/ionization-MS platform, which employs droplet microfluidics for onplate nanoliter reactions. A chromatographic separation of a proteolytic digest is deposited at a high frequency on the microarray. In this way, a short separation run is archived into thousands of nanoliter reaction cavities, and chromatographic peaks are spread over multiple array spots. After fractionation, each other spot is treated with PNGaseF to generate two correlated traces within one run, one with treated spots where glycans are enzymatically released from the peptides, and one containing the intact glycopeptides. Mining for distinct glycosites is performed by searching for the predicted deglycosylated peptides in the treated trace. An identified peptide then leads directly to the position of the "intact" glycopeptide clusters, which are located in the adjacent spots. Furthermore, the deglycosylated peptide can be sequenced efficiently in a simple collision-induced dissociation-MS experiment. We applied the microarray approach to a detailed site-specific glycosylation analysis of human serum IgM. By scanning the treated spots with low-resolution matrix assisted laser desorption/ionization-time-offlight-MS, we observed all five deglycosylated peptides, including the one originating from the secretory chain. A detailed glycopeptide characterization was then accomplished on the adjacent, untreated spots with high mass resolution and high mass accuracy using a matrix assisted laser desorption ionization-Fourier transform-MS. We present the first detailed and comprehensive mass spectrometric analysis on the glycopeptide level for human polyclonal IgM with high mass accuracy. Besides complex type glycans on Asn 395, 332, 171, and on the J chain, we observed oligomannosidic glycans on Asn 563, Asn 402 and minor amounts of oligomannosidic glycans on the glycosite Asn 171. Furthermore, hybrid type glycans were found on Asn 402, Asn 171 and in traces Asn 332. Molecular & Cellular

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confidence: 99%

A Microarray-Matrix-assisted Laser Desorption/Ionization-Mass Spectrometry Approach for Site-specific Protein N-glycosylation Analysis, as Demonstrated for Human Serum Immunoglobulin M (IgM)*

Pabst

Kuster

Wahl³

et al. 2015

Molecular & Cellular Proteomics

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“…For focused glycoproteomics, several techniques using lectins or antibodies enabling the large-scale identification of glycoproteins have recently been developed (17)(18)(19). Notably, Zhang et al reported a method for the selective isolation of peptides based on chemical oxidation of the carbohydrate moiety and subsequent conjugation to a solid support using hydrazide chemistry (20 -26).…”

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confidence: 99%

Sialic Acid-focused Quantitative Mouse Serum Glycoproteomics by Multiple Reaction Monitoring Assay

Kurogochi

Matsushista

Amano

et al. 2010

Molecular & Cellular Proteomics

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Clinical proteomics focusing on the identification and validation of biomarkers and the discovery of proteins as therapeutic targets is an emerging and highly important area of proteomics. Biomarkers are measurable indicators of a specific biological state (particularly one relevant to the risk of contraction) and the presence or the stage of disease, and are thus expected to be useful for the prediction, detection, and diagnosis of disease as well as to follow the efficacy, toxicology, and side effects of drug treatment, and to provide new functional insights into biological processes.At present, proteomics methods based on mass spectrometry (MS) have emerged as the preferred strategy for discovery of diagnostic, prognostic, and therapeutic protein biomarkers. Most biomarker discovery studies use unbiased, "identified-based" approaches that rely on high performance mass spectrometers and extensive sample processing. Semiquantitative comparisons of protein relative abundance between disease and control patient samples are used to identify proteins that are differentially expressed and, thus, to populate lists of potential biomarkers. De novo proteomics discovery experiments often result in tens to hundreds of candidate biomarkers that must be subsequently verified in serum. However, despite the large numbers of putative biomarkers, only a small number of them are passed through the development and validation process into clinical practice, and their rate of introduction is declining. The first non-standard abbreviation (MS above is standard) must be footnoted the same as the abbreviation footnote, and MRM must be the first abbreviation in the list because it is the one footnoted. After that the order does not matter.Targeted proteomics using multiple reaction monitoring (MRM) 1 is emerging as a technology that complements the discovery capabilities of shotgun strategies as well as an alternative powerful novel MS-based approach to measure a series of candidate biomarkers (1-7). Therefore, MRM is expected to provide a powerful high throughput platform for biomarker validation, although clinical validation of novel biomarkers has been traditionally relying on immunoassays (8, 9). MRM exploits the unique capabilities of triple quadrupoles (QQQ) MS for quantitative analysis. In MRM, the first and the third quadrupoles act as filters to specifically select predefined m/z values corresponding to the peptide precursor ion and specific fragment ion of the peptide, whereas the second quadrupole serves as collision cell. Several such transitions (precursor/fragment ion pairs) are monitored over time, yielding a set of chromatographic traces with retention time and signal intensity for a specific transition as coordinates. These measurements have been multiplexed to provide 30 or From the

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“…Over the past decade, various glycopeptide desalting methodologies have been developed including dialysis, gel filtration and LC. Among these, reverse phase LC C18 materials are commonly used to desalt glycopeptides 12,13 because of their high binding capacity for most biological molecules. C18-based materials can remove the low molecular weight contaminants, improving the glycopeptide intensity.…”

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confidence: 99%

Click maltose as an alternative to reverse phase material for desalting glycopeptides

Guo

et al. 2011

Analyst

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Desalting peptides before mass spectrometry analysis is important because salts lead to adduct formation, increased chemical noise and ion suppression effect. A high concentration of salt can clog nanoelectrospray ionization (ESI) emitters. The reverse phase C18 material is commonly used to desalt peptides because of its high binding capacity. However, peptides with high hydrophilicity, such as glycopeptides, are not retained well on this material, resulting in the loss of peptide information. To improve the efficiency of glycopeptide desalting, we introduced a hydrophilic interaction chromatography (HILIC)-based material named click maltose. Four glycoproteins, horseradish peroxidase (HRP), human serum immunoglobulin G (IgG), bovine ribonuclease B (RNase B), and a-1 acid glycoprotein (AGP) were chosen as models and their glycopeptides were desalted with click maltose, AQ C18, Empore C18 and ZipTip C18. Click maltose as a HILIC material exhibited better performance than the other three C18 materials for both number of targeted glycopeptides and their corresponding intensities. In addition, accurate glycopeptide profiling was achieved with click maltose desalting regardless of peptide lengths and glycan types.

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Assessment of lectin and HILIC based enrichment protocols for characterization of serum glycoproteins by mass spectrometry

Cited by 122 publications

References 49 publications

A Microarray-Matrix-assisted Laser Desorption/Ionization-Mass Spectrometry Approach for Site-specific Protein N-glycosylation Analysis, as Demonstrated for Human Serum Immunoglobulin M (IgM)*

A Microarray-Matrix-assisted Laser Desorption/Ionization-Mass Spectrometry Approach for Site-specific Protein N-glycosylation Analysis, as Demonstrated for Human Serum Immunoglobulin M (IgM)*

Sialic Acid-focused Quantitative Mouse Serum Glycoproteomics by Multiple Reaction Monitoring Assay

Click maltose as an alternative to reverse phase material for desalting glycopeptides

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