A Novel Spectral Annotation Strategy Streamlines Reporting of Mono-ADP-ribosylated Peptides Derived from Mouse Liver and Spleen in Response to IFN-γ.

Autor:	Kuraoka S; Center for Interdisciplinary Cardiovascular Sciences, Division of Cardiovascular Medicine, Department of Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA., Higashi H; Center for Interdisciplinary Cardiovascular Sciences, Division of Cardiovascular Medicine, Department of Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA., Yanagihara Y; Center for Interdisciplinary Cardiovascular Sciences, Division of Cardiovascular Medicine, Department of Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA., Sonawane AR; Center for Interdisciplinary Cardiovascular Sciences, Division of Cardiovascular Medicine, Department of Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA; Center for Excellence in Vascular Biology, Division of Cardiovascular Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA., Mukai S; Center for Interdisciplinary Cardiovascular Sciences, Division of Cardiovascular Medicine, Department of Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA., Mlynarchik AK; Center for Interdisciplinary Cardiovascular Sciences, Division of Cardiovascular Medicine, Department of Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA., Whelan MC; Center for Interdisciplinary Cardiovascular Sciences, Division of Cardiovascular Medicine, Department of Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA., Hottiger MO; Department of Molecular Mechanisms of Disease, University of Zurich, Zurich, Switzerland., Nasir W; Thermo Fisher Scientific (Bremen) GmbH, Bremen, Germany., Delanghe B; Thermo Fisher Scientific (Bremen) GmbH, Bremen, Germany., Aikawa M; Center for Interdisciplinary Cardiovascular Sciences, Division of Cardiovascular Medicine, Department of Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA; Center for Excellence in Vascular Biology, Division of Cardiovascular Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA; Channing Division of Network Medicine, Department of Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA. Electronic address: maikawa@bwh.harvard.edu., Singh SA; Center for Interdisciplinary Cardiovascular Sciences, Division of Cardiovascular Medicine, Department of Medicine, Brigham Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA. Electronic address: sasingh@bwh.harvard.edu.
Jazyk:	angličtina
Zdroj:	Molecular & cellular proteomics : MCP [Mol Cell Proteomics] 2022 Apr; Vol. 21 (4), pp. 100153. Date of Electronic Publication: 2021 Sep 28.
DOI:	10.1016/j.mcpro.2021.100153
Abstrakt:	Mass-spectrometry-enabled ADP-ribosylation workflows are developing rapidly, providing researchers a variety of ADP-ribosylome enrichment strategies and mass spectrometric acquisition options. Despite the growth spurt in upstream technologies, systematic ADP-ribosyl (ADPr) peptide mass spectral annotation methods are lacking. HCD-dependent ADP-ribosylome studies are common, but the resulting MS2 spectra are complex, owing to a mixture of b/y-ions and the m/p-ion peaks representing one or more dissociation events of the ADPr moiety (m-ion) and peptide (p-ion). In particular, p-ions that dissociate further into one or more fragment ions can dominate HCD spectra but are not recognized by standard spectral annotation workflows. As a result, annotation strategies that are solely reliant upon the b/y-ions result in lower spectral scores that in turn reduce the number of reportable ADPr peptides. To improve the confidence of spectral assignments, we implemented an ADPr peptide annotation and scoring strategy. All MS2 spectra are scored for the ADPr m-ions, but once spectra are assigned as an ADPr peptide, they are further annotated and scored for the p-ions. We implemented this novel workflow to ADPr peptides enriched from the liver and spleen isolated from mice post 4 h exposure to systemic IFN-γ. HCD collision energy experiments were first performed on the Orbitrap Fusion Lumos and the Q Exactive, with notable ADPr peptide dissociation properties verified with CID (Lumos). The m-ion and p-ion series score distributions revealed that ADPr peptide dissociation properties vary markedly between instruments and within instrument collision energy settings, with consequences on ADPr peptide reporting and amino acid localization. Consequentially, we increased the number of reportable ADPr peptides by 25% (liver) and 17% (spleen) by validation and the inclusion of lower confidence ADPr peptide spectra. This systematic annotation strategy will streamline future reporting of ADPr peptides that have been sequenced using any HCD/CID-based method. Competing Interests: Conflict of interest B. D. and W. N. are employees of Thermo Fisher Scientific. All other authors declare no competing interests. (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
Externí odkaz:	Zobrazit plný text záznamu