Cole, J. orcid.org/0000-0002-5258-5835, Hanson, E.J., James, D.C. et al. (2 more authors) (2019) Comparison of data acquisition methods for the identification and quantification of histone post-translational modifications on a Q Exactive HF hybrid quadrupole Orbitrap mass spectrometer. Rapid Communications in Mass Spectrometry, 33 (10). pp. 897-906. ISSN 0951-4198
Abstract
RATIONALE: Histone PTMs play key roles in regulating eukaryotic gene expression. Mass spectrometry (MS) has emerged as a powerful method to characterize and quantify histone PTMs as it allows unbiased identification and quantification of multiple histone PTMs including combinations of the modifications present. METHODS: In this study we compared a range of data acquisition methods for the identification and quantification of the histone PTMs using a Q Exactive HF Orbitrap. We compared three different data-dependent analysis (DDA) methods with MS2 resolutions of 120K, 60K, 30K. We also compared a range of data-independent analysis (DIA) methods using MS2 isolation windows of 20 m/z and DIAvw to identify and quantify histone PTMs in Chinese Hamster Ovary (CHO) cells. RESULTS: The increased number of MS2 scans afforded by the lower resolution methods resulted in a higher number of queries, peptide sequence matches (PSMs) and a higher number of peptide proteoforms with a Mascot Ion score greater than 46. No difference in the proportion of peptide proteoforms with Delta scores >17 was observed. Comparing the data acquisition methods increased repeatability in terms of lower CVs afforded by DIA MS1 60K MS2 30K 20m/z isolation windows was observed. CONCLUSION: We observed that DIA which offers advantages in flexibility and identification of isobaric peptide proteoforms performs as well as DDA in the analysis of histone PTMs. We were able to identify 71 modified histone peptides for histone H3 and H4 and quantified 64 across each of the different acquisition methods.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2019 Wiley. This is an author produced version of a paper subsequently published in Rapid Communications in Mass Spectrometry. Uploaded in accordance with the publisher's self-archiving policy. |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Engineering (Sheffield) > Department of Chemical and Biological Engineering (Sheffield) The University of Sheffield > Sheffield Teaching Hospitals |
Funding Information: | Funder Grant number BIOTECHNOLOGY AND BIOLOGICAL SCIENCES RESEARCH COUNCIL (BBSRC) BB/M012166/1 The Wellcome Trust 104437/Z/14/Z |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 10 Apr 2019 10:46 |
Last Modified: | 23 Nov 2021 10:27 |
Status: | Published |
Publisher: | Wiley |
Refereed: | Yes |
Identification Number: | 10.1002/rcm.8401 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:144743 |