IR-Based Protein & Peptide Quantitation

Despite the successful application of tandem mass tags (TMT) for peptide quantitation, missing reporter ions in higher energy collisional dissociation (HCD) spectra remains a challenge for consistent quantitation, especially for peptides with labile post-translational modifications. Ultraviolet photodissociation (UVPD) is an alternative ion activation method shown to provide superior coverage for sequencing of peptides and intact proteins. Here, we optimized and evaluated 193 nm UVPD for the characterization of TMT-labeled model peptides, HeLa proteome, and N-glycopeptides from model proteins. UVPD yielded the same TMT reporter ions as HCD, at m/z 126–131. Additionally, UVPD produced a wide range of fragments that yielded more complete characterization of glycopeptides and less frequent missing TMT reporter ion channels, whereas HCD yielded a strong tradeoff between characterization and quantitation of TMT-labeled glycopeptides. However, the lower fragmentation efficiency of UVPD yielded fewer peptide identifications than HCD. Overall, 193 nm UVPD is a valuable tool that provides an alternative to HCD for the quantitation of large and highly modified peptides with labile PTMs. Continued development of instrumentation specific to UVPD will yield greater fragmentation efficiency and fulfil the potential of UVPD to be an all-in-one spectrum ion activation method for broad use in the field of proteomics.

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Peptide quantitation using primary amine selective metal chelation labels for mass spectrometry

Chemical Communications ◽

10.1039/b926290c ◽

2010 ◽

Vol 46 (30) ◽

pp. 5479 ◽

Cited By ~ 12

Author(s):

Thomas J. Kerr ◽

John A. McLean

Keyword(s):

Mass Spectrometry ◽

Primary Amine ◽

Metal Chelation ◽

Peptide Quantitation

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Development of a UPLC-IDA-ICP-MS/MS method for peptide quantitation in plasma by Se-labelling, and comparison to S-detection of the native peptide

Journal of Analytical Atomic Spectrometry ◽

10.1039/c8ja00341f ◽

2019 ◽

Vol 34 (2) ◽

pp. 375-383 ◽

Cited By ~ 3

Author(s):

Freja Grønbæk-Thorsen ◽

Stefan Stürup ◽

Bente Gammelgaard ◽

Laura Hyrup Møller

Keyword(s):

Native Peptide ◽

Icp Ms ◽

Peptide Quantitation

Se-labelling improves quantitation of peptides in plasma significantly compared to S-detection of the native peptide.

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Solid-phase peptide quantitation assay using labeled monoclonal antibody and glutaraldehyde fixation

Analytical Biochemistry ◽

10.1016/0003-2697(88)90539-8 ◽

1988 ◽

Vol 174 (1) ◽

pp. 224-234 ◽

Cited By ~ 10

Author(s):

Philip G. Kasprzyk ◽

Frank Cuttitta ◽

Ingalill Avis ◽

Yoichi Nakanishi ◽

Anthony Treston ◽

...

Keyword(s):

Monoclonal Antibody ◽

Solid Phase ◽

Glutaraldehyde Fixation ◽

Peptide Quantitation

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Protein Expression Analysis: From ‘Tip of the Iceberg’ to a Global Method

Disease Markers ◽

10.1155/2001/385075 ◽

2001 ◽

Vol 17 (4) ◽

pp. 235-246 ◽

Cited By ~ 3

Author(s):

Peter James

Keyword(s):

Protein Expression ◽

Expression Analysis ◽

Spectrometric Analysis ◽

Two Dimensional ◽

Two Dimensional Gel Electrophoresis ◽

Mass Spectrometers ◽

Global Method ◽

Stable Isotopic ◽

Highly Effective ◽

Peptide Quantitation

In this review I will describe the advances that have recently been made in ‘traditional’ two-dimensional gel based protein expression analysis. A major jump has been made toward the automation of gel image analysis and comparison, one of the major bottlenecks in the analysis chain as well as the automation of spot excision and preparation for mass spectrometric analysis. Currently the gel-based ‘proteome mapping’ approach is highly effective and 300 gels and over 10,000 spots a week can be analysed. Very recently, viable alternatives to the use of two-dimensional gel electrophoresis have emerged and these approaches are discussed here. In combination with the recently developed stable isotopic tagging methods for peptide quantitation and new mass spectrometers, this emerging technology will be a rapid and highly effective alternative to gel-based methods with few of the latter's shortcomings.

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Proteomics reveals that quinoa bioester promotes replenishing effects in epidermal tissue

Scientific Reports ◽

10.1038/s41598-020-76325-6 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Amanda C. Camillo-Andrade ◽

Marlon D. M. Santos ◽

Juliana S. G. Fischer ◽

Bruna B. Swinka ◽

Bruna Bosquetti ◽

...

Keyword(s):

Tissue Regeneration ◽

Shotgun Proteomics ◽

Reversed Phase ◽

Premature Aging ◽

Analysis Tool ◽

Epidermal Tissue ◽

Age Related ◽

Peptide Quantitation ◽

Data Analysis Tool ◽

Significant Attention

Abstract The continuous search for natural products that attenuate age-related losses has increasingly gained notice; among them, those applicable for skin care have drawn significant attention. The bioester generated from the Chenopodium quinoa’s oil is a natural-origin ingredient described to produce replenishing skin effects. With this as motivation, we used shotgun proteomics to study the effects of quinoa bioester on human reconstructed epidermis tridimensional cell cultures after 0, 3, 6, 12, 24, and 48 h of exposure. Our experimental setup employed reversed-phase nano-chromatography coupled online with an Orbitrap-XL and PatternLab for proteomics as the data analysis tool. Extracted ion chromatograms were obtained as surrogates for relative peptide quantitation. Our findings spotlight proteins with increased abundance, as compared to the untreated cell culture counterparts at the same timepoints, that were related to preventing premature aging, homeostasis, tissue regeneration, protection against ultraviolet radiation and oxidative damage.

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