Proteome Letters
Online ISSN : 2432-2776
ISSN-L : 2432-2776
Volume 1, Issue 2
Displaying 1-6 of 6 articles from this issue
  • Junichi Kamiie, Naoyuki Aihara, Kinji Shirota
    2016 Volume 1 Issue 2 Pages 57-62
    Published: 2016
    Released on J-STAGE: May 21, 2018
    JOURNAL FREE ACCESS

    Selected reaction monitoring mode (SRM)-based strategies have recently been developed for quantitation of proteins. Using sequence information from the proteins, these strategies can be used to develop simultaneous quantitative methods for several proteins of interest. In this article, we discussed the problems and potential roles of SRM-based strategies in protein quantitation. “Absolute” quantitation of proteins is one of the strengths of SRM strategies. For mass spectrometry, protein samples should be digested with enzymes to their component peptides. But we cannot correct for the sample loss or efficiency of enzyme digestion in this preparatory step. Therefore, quantitative values from SRM strategies may underestimate protein abundance. In this article, we introduced our solution to this problem with isotope-labeled protein as quantitative reference. Also, we have developed a PTM profiling method with isotope-labeled full-length proteins as references. This method makes it possible to obtain the modification sites and rates of proteins using differential analysis for the quantitative values of each peptide.

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  • Akiyasu C. Yoshizawa
    2016 Volume 1 Issue 2 Pages 63-80
    Published: 2016
    Released on J-STAGE: May 21, 2018
    JOURNAL FREE ACCESS

    For mass spectrometry based-proteomics studies, computational analyses of obtained data are indispensable. However, the analysis methodologies and software for mass spectrometry and/or proteomics are currently still under development and many problems thus remain unfixed; consequently, researchers, especially experimental scientists, often suffer from technical issues and popular misinterpretations. Based on these problems, we describe in this review the computational processes for protein identification for proteomics beginners, especially the algorithms of database search and related basic issues: the comparison of de novo sequencing method and database search method, the effects of PTM detection on the search results, an overview of life science databases, and tips and cautions for their application to database searches.

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  • Noriaki Arakawa, Hisashi Hirano
    2016 Volume 1 Issue 2 Pages 81-87
    Published: 2016
    Released on J-STAGE: May 21, 2018
    JOURNAL FREE ACCESS

    Cancer is a leading cause of death. Although recent advances in proteomics technologies have inspired hope that new blood-based biomarkers for early cancer detection will be discovered, few have been introduced to the clinic. Biomarker discovery using plasma or serum is challenging. Detecting cancer cell–derived proteins is extremely difficult in highly complex biological fluids that include an abundance of proteins produced from the liver. On the other hand, a conditioned medium of cultured cancer cells containing an abundance of proteins secreted or released from the cells and those shed from the cell surface, which is referred to as the “secretome”, is a promising source of biomarker discovery. However, obstacles in secretome analysis include sample preparation, effective selection of biomarker candidates, and assay system development. Herein we introduce findings determined via secretome analysis with an emphasis on methodologies developed for biomarker discovery based on culture cell lines and their advantages and disadvantages.

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  • Fumio Matsuda
    2016 Volume 1 Issue 2 Pages 89-94
    Published: 2016
    Released on J-STAGE: May 21, 2018
    JOURNAL FREE ACCESS

    Targeted proteome analysis is an approach for quantitative analysis of multiple target peptides using liquid chromatography-triple quadrupole mass spectrometry. Although it is essentially similar to that of quantitative analysis of small molecules, several specific techniques are required for targeted proteome analysis including sample preparation methods, handling of nano-LC, and construction of SRM assay methods. Recent progresses in the sample preparation protocols, improved stability of modern nanoLC machines, and development of softwares enable a wide application of the targeted proteome analysis in biological studies.

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  • Takeshi Masuda, Yasushi Ishihama
    2016 Volume 1 Issue 2 Pages 95-100
    Published: 2016
    Released on J-STAGE: May 21, 2018
    JOURNAL FREE ACCESS

    We have reported a phase transfer surfactants (PTSs)-aided sample preparation protocol for shotgun proteomics. The PTS protocol using deoxycholate and lauroylsarcosin increases the solubility of membrane proteins and enhances trypsin and lysyl endopeptidase activities. The PTSs become more hydrophobic at acidic pH. Based on this property, PTSs are removable from sample solutions by the liquid-liquid extraction method with ethyl acetate as organic solvent, while peptides are remained in the aqueous phase. Compared with protocols using a RapiGest or a filter–aided sample preparation protocol, the number of identified proteins and the recovery of peptides are significantly increased in the PTS protocol, suggesting that the PTSs protocol is more effective for the large-scale unbiased proteomics than these protocols.

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  • Masaki Matsumoto
    2016 Volume 1 Issue 2 Pages 101-106
    Published: 2016
    Released on J-STAGE: May 21, 2018
    JOURNAL FREE ACCESS

    Advances in sensitivity, scanning speed, and resolution of mass spectrometry extensively facilitated comprehensive identification and quantification of proteome. However, one of the major techniques used in the current proteomics, data-dependent acquisition (DDA), has technical limitations that prevent in-depth proteome analysis across large number of samples. Recently, targeted proteomics, such as multiple-reaction monitoring (MRM) has been developed to promote high-throughput, accurate and consistent quantification of given protein sets, that serves the extended application of proteomics for biology. In this review, I describe the current topics in targeted proteomics and the sample processing procedure required for targeted proteomics.

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