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PXD025971

PXD025971 is an original dataset announced via ProteomeXchange.

Dataset Summary
TitleRapid Highly-Efficient Digestion and Peptide Mapping of Adeno-Associated Viruses
Description[15:18] Toole, Estee Adeno-Associated Viruses (AAVs) comprise an area of rapidly growing interest due to their ability to act as a gene delivery vehicle in novel gene therapy strategies and vaccine development. Peptide mapping is a common technique in the biopharmaceutical industry to confirm the correct sequence, product purity, PTMs, and stability. However, conventional peptide mapping is time-consuming and has proven difficult to reproduce with viral capsids because of their high structural stability and the suboptimal localization of trypsin cleavage sites in the AAV protein sequences. In this study, we present an optimized workflow that provides thorough characterization within one day. This workflow is also highly reproducible due to its simplicity having very few steps, and easy to perform proteolytic digestion utilizing thermally-stable pepsin, active at 70°C in acidic conditions. The acidic conditions of the peptic digestions drive viral capsid denaturation and improve cleavage site accessibility. We characterized the efficiency and ease of digestion through peptide mapping of the AAV2 viral capsid protein. Using nanoflow liquid chromatography coupled with tandem mass spectrometry, we achieved 100% sequence coverage of the low abundance VP1 capsid protein with a digestion process taking only 10 minutes to prepare and 30 minutes to complete the digestion.
HostingRepositoryPRIDE
AnnounceDate2022-02-17
AnnouncementXMLSubmission_2022-02-17_03:28:08.254.xml
DigitalObjectIdentifier
ReviewLevelPeer-reviewed dataset
DatasetOriginOriginal dataset
RepositorySupportUnsupported dataset by repository
PrimarySubmitterAndrew Williamson
SpeciesList scientific name: Adeno-associated virus - 2; NCBI TaxID: 10804;
ModificationListmonohydroxylated residue; deamidated residue
InstrumentQ Exactive
Dataset History
RevisionDatetimeStatusChangeLog Entry
02021-05-12 01:46:22ID requested
12022-02-17 03:28:09announced
Publication List
Toole EN, Dufresne C, Ray S, Schwann A, Cook K, Ivanov AR, Rapid Highly-Efficient Digestion and Peptide Mapping of Adeno-Associated Viruses. Anal Chem, 93(30):10403-10410(2021) [pubmed]
Keyword List
submitter keyword: [15:18] Toole, Estee Gene Therapy, AAV, Viral Therapies, Peptide Mapping, Pepsin, Trypsin, nanoLC, proteolysis
Contact List
Estee Toole
contact affiliationThermo Fisher Scientific
contact emailestee.toole@thermofisher.com
lab head
Andrew Williamson
contact affiliationThermo Fisher Scientific
contact emailandrew.williamson@thermofisher.com
dataset submitter
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Dataset FTP location
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