title: Adenovirus composition, proteolysis, and disassembly studied by in-depth qualitative and quantitative proteomics.
abstract: Using high-resolution MS-based proteomics in combination with multiple protease digestion, we profiled, with on average 90% sequence coverage, all 13 viral proteins present in an human adenovirus (HAdV) vector. This in-depth profile provided multiple peptide-based evidence on intrinsic protease activity affecting several HAdV proteins. Next, the generated peptide library was used to develop a targeted proteomics method using selected reaction monitoring (SRM) aimed at quantitative profiling of the stoichiometry of all 13 proteins present in the HAdV. We also used this method to probe the release of specific virus proteins initiated by thermal stimulation, mimicking the early stage of HAdV disassembly during entry into host cells. We confirmed the copy numbers of the most well characterized viral capsid components and established the copy numbers for proteins whose stoichiometry has so far not been accurately defined. We also found that heating HAdV induces the complete release of the penton base and fiber proteins as well as a substantial release of protein VIII and VI. For these latter proteins, maturational proteolysis by the adenoviral protease leads to the differential release of fragments with certain peptides being fully released and others largely retained in the AdV particles. This information is likely to be beneficial for the ongoing interpretation of high resolution cryoEM and x-ray electron density maps.
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Centre for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Padualaan 8, 3584 CH Utrecht, The Netherlands,; Netherlands Proteomics Center, Padualaan 8, 3584 CH, Utrecht, The Netherlands.
authors.forename: Marco
authors.initials: M
authors.lastname: Benevento
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Centre for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Padualaan 8, 3584 CH Utrecht, The Netherlands,; Netherlands Proteomics Center, Padualaan 8, 3584 CH, Utrecht, The Netherlands.
authors.forename: Serena
authors.initials: S
authors.lastname: Di Palma
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Centre for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Padualaan 8, 3584 CH Utrecht, The Netherlands,; Netherlands Proteomics Center, Padualaan 8, 3584 CH, Utrecht, The Netherlands.
authors.forename: Joost
authors.initials: J
authors.lastname: Snijder
authors.affiliation: Department of Immunology and Microbial Science, The Scripps Research Institute, La Jolla, California 92037, and.
authors.forename: Crystal L
authors.initials: CL
authors.lastname: Moyer
authors.affiliation: Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, California 92037.
authors.forename: Vijay S
authors.initials: VS
authors.lastname: Reddy
authors.affiliation: Department of Immunology and Microbial Science, The Scripps Research Institute, La Jolla, California 92037, and.
authors.forename: Glen R
authors.initials: GR
authors.lastname: Nemerow
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Centre for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Padualaan 8, 3584 CH Utrecht, The Netherlands,; Netherlands Proteomics Center, Padualaan 8, 3584 CH, Utrecht, The Netherlands,. Electronic address: A.J.R.Heck@uu.nl.
authors.forename: Albert J R
authors.initials: AJR
authors.lastname: Heck
mesh_terms: D000260:Adenoviruses, Human / Q000502:physiology* / Q000648:ultrastructure; D002460:Cell Line; D006801:Humans; D040901:Proteomics; D014764:Viral Proteins / Q000378:metabolism*; D019065:Virus Assembly / Q000502:physiology*
chemical_list: D014764:Viral Proteins
keywords: Adenovirus; Mass Spectrometry (MS); Proteomics; Virus Entry; Virus Structure