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Recent Progress in FD-LC-MS/MS Proteomics Method
Through the course of our bio-analytical chemistry studies, we developed a novel proteomics analysis method, FD-LC-MS/MS (fluorogenic derivatization-liquid chromatography-tandem mass spectrometry). This method consists of fluorogenic derivatization (FD), LC separation, and detection/quantification o...
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Frontiers Media S.A.
2021
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8220812/ https://www.ncbi.nlm.nih.gov/pubmed/34178939 http://dx.doi.org/10.3389/fchem.2021.640336 |
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author | Kobayashi, Hiroshi Imai, Kazuhiro |
author_facet | Kobayashi, Hiroshi Imai, Kazuhiro |
author_sort | Kobayashi, Hiroshi |
collection | PubMed |
description | Through the course of our bio-analytical chemistry studies, we developed a novel proteomics analysis method, FD-LC-MS/MS (fluorogenic derivatization-liquid chromatography-tandem mass spectrometry). This method consists of fluorogenic derivatization (FD), LC separation, and detection/quantification of the derivatized proteins, followed by isolation, tryptic digestion of the isolated proteins, and final identification of the isolated proteins using electrospray ionization nano-LC-MS/MS of the generated peptide mixtures with a probability-based protein identification algorithm. In this review, we will present various examples where this method has been used successfully to identify expressed proteins in individual human cells. FD-LC-MS/MS is also suitable for differential proteomics analysis. Here, two biological samples are treated by the same steps mentioned above, and the two chromatograms obtained are compared to identify peaks with different intensities (variation in protein levels). Associated peak fractions are then isolated, and the differentially expressed proteins between the two samples are identified by LC-MS/MS. Several biomarkers for cancers have been identified by FD-LC-MS/MS. For more efficient separation, nano-flow LC with a phenyl-bonded monolithic silica-based capillary column was adopted for cell-expressed intact protein analysis. The derivatized human cell proteins (K562) and yeast cell (Saccharomyces cerevisiae) proteins as model intact cell proteins were analyzed by nano-flow LC with fluorescence detection. More than 1,300 protein peaks were separated/detected from both cells. For straightforward comparison of multiple peak separation profiles, a novel type of chromatogram display, termed the “spiderweb” chromatogram, was developed. A nano-LC-FD-LC-mass spectrometry trial for molecular weight estimation of FD proteins has also been conducted. |
format | Online Article Text |
id | pubmed-8220812 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-82208122021-06-24 Recent Progress in FD-LC-MS/MS Proteomics Method Kobayashi, Hiroshi Imai, Kazuhiro Front Chem Chemistry Through the course of our bio-analytical chemistry studies, we developed a novel proteomics analysis method, FD-LC-MS/MS (fluorogenic derivatization-liquid chromatography-tandem mass spectrometry). This method consists of fluorogenic derivatization (FD), LC separation, and detection/quantification of the derivatized proteins, followed by isolation, tryptic digestion of the isolated proteins, and final identification of the isolated proteins using electrospray ionization nano-LC-MS/MS of the generated peptide mixtures with a probability-based protein identification algorithm. In this review, we will present various examples where this method has been used successfully to identify expressed proteins in individual human cells. FD-LC-MS/MS is also suitable for differential proteomics analysis. Here, two biological samples are treated by the same steps mentioned above, and the two chromatograms obtained are compared to identify peaks with different intensities (variation in protein levels). Associated peak fractions are then isolated, and the differentially expressed proteins between the two samples are identified by LC-MS/MS. Several biomarkers for cancers have been identified by FD-LC-MS/MS. For more efficient separation, nano-flow LC with a phenyl-bonded monolithic silica-based capillary column was adopted for cell-expressed intact protein analysis. The derivatized human cell proteins (K562) and yeast cell (Saccharomyces cerevisiae) proteins as model intact cell proteins were analyzed by nano-flow LC with fluorescence detection. More than 1,300 protein peaks were separated/detected from both cells. For straightforward comparison of multiple peak separation profiles, a novel type of chromatogram display, termed the “spiderweb” chromatogram, was developed. A nano-LC-FD-LC-mass spectrometry trial for molecular weight estimation of FD proteins has also been conducted. Frontiers Media S.A. 2021-06-04 /pmc/articles/PMC8220812/ /pubmed/34178939 http://dx.doi.org/10.3389/fchem.2021.640336 Text en Copyright © 2021 Kobayashi and Imai. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Chemistry Kobayashi, Hiroshi Imai, Kazuhiro Recent Progress in FD-LC-MS/MS Proteomics Method |
title | Recent Progress in FD-LC-MS/MS Proteomics Method |
title_full | Recent Progress in FD-LC-MS/MS Proteomics Method |
title_fullStr | Recent Progress in FD-LC-MS/MS Proteomics Method |
title_full_unstemmed | Recent Progress in FD-LC-MS/MS Proteomics Method |
title_short | Recent Progress in FD-LC-MS/MS Proteomics Method |
title_sort | recent progress in fd-lc-ms/ms proteomics method |
topic | Chemistry |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8220812/ https://www.ncbi.nlm.nih.gov/pubmed/34178939 http://dx.doi.org/10.3389/fchem.2021.640336 |
work_keys_str_mv | AT kobayashihiroshi recentprogressinfdlcmsmsproteomicsmethod AT imaikazuhiro recentprogressinfdlcmsmsproteomicsmethod |