dELTA-MS: A Mass Spectrometry-Based Proteomics Approach for Identifying ADP-Ribosylation Sites and Forms
Journal of Proteome Research, vol. 24, pp. 1791–1803
Abstract
ADP-ribosylation, characterized by the addition of adenosine diphosphate ribose, can occur in both monomeric (MARylation) and polymeric (PARylation) forms. Little is known about the specific contributions of MARylation and PARylation to cellular processes due to a lack of tools for jointly investigating these individual forms. We present a novel mass spectrometry (MS)-based proteomics approach that preserves information about the native ADP-ribosylation form associated with the modification site within a single proteomics experiment. Our workflow enables the simplified, binary identification of ADP-ribosylation forms, avoiding some challenges typically presented by PARylated peptides during MS analysis. Our method uses the coronaviral enzyme NS2 to reverse our previous labeling approach, ELTA, which enzymatically labels the terminal ADP-ribose. NS2 deconjugates ELTA-labeled free or peptide-conjugated ADP-ribose monomers and polymers (thereby termed dELTA), leaving behind a signature phosphate. Our dELTA-MS workflow involves ELTA labeling, dELTA deconjugation, and further processing using Deinococcus radiodurans poly(ADP-ribose) glycohydrolase (DrPARG), resulting in two distinct mass shifts for MARylation and PARylation sites. We demonstrate the feasibility of this workflow for proteomics analyses using proof-of-principle peptide standards. dELTA-MS thus creates possibilities to reveal the fundamental biology of ADP-ribosylation and explore its dysregulation, in terms of both sites and forms, associated with disease progression.
Authors 14
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Affiliation as printed
Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health
Johns Hopkins University
Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, Maryland 21205, United States
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Washington University in St. Louis
Affiliation as printed
Department of Biochemistry and Molecular Biophysics
Washington University School of Medicine
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, Missouri 63110, United States
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Washington University in St. Louis
Affiliation as printed
Department of Biochemistry and Molecular Biophysics
Washington University School of Medicine
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, Missouri 63110, United States
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Affiliation as printed
Department of Neurology
Institute for Cell Engineering Johns Hopkins University
Department of Neurology, Institute for Cell Engineering Johns Hopkins University, Baltimore, Maryland 21205, United States
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Affiliation as printed
Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health
Johns Hopkins University
Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, Maryland 21205, United States
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Affiliation as printed
Gorlaeus Laboratories, Leiden Institute of Chemistry, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands
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Affiliation as printed
EpiCypher Inc
EpiCypher Inc., Durham, North Carolina 27709, United States
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Affiliation as printed
EpiCypher Inc
EpiCypher Inc., Durham, North Carolina 27709, United States
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The University of Texas Southwestern Medical Center
Affiliation as printed
Department of Biochemistry
University of Texas Southwestern Medical Center
Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas 75390, United States
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Affiliation as printed
Gorlaeus Laboratories, Leiden Institute of Chemistry, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands
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Affiliation as printed
Department of Neurology
Institute for Cell Engineering Johns Hopkins University
Department of Neurology, Institute for Cell Engineering Johns Hopkins University, Baltimore, Maryland 21205, United States
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Washington University in St. Louis
Affiliation as printed
Department of Biochemistry and Molecular Biophysics
Washington University School of Medicine
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, Missouri 63110, United States
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Benjamin C. Orsburn corresponding
Affiliation as printed
Department of Pharmacology and Chemical Biology
University of Pittsburgh
Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, Pennsylvania 15260, United States
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Anthony K. L. Leung corresponding
Affiliation as printed
Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health
Department of Genetic Medicine, School of Medicine
Department of Molecular Biology and Genetics, School of Medicine
Johns Hopkins University
Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, Maryland 21205, United States
Department of Genetic Medicine, School of Medicine, Johns Hopkins University, Baltimore, Maryland 21205, United States
Department of Molecular Biology and Genetics, School of Medicine, Johns Hopkins University, Baltimore, Maryland 21205, United States
Department of Oncology, School of Medicine, Johns Hopkins University, Baltimore, Maryland 21205, United States
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