PXD030094-1

PXD030094 is an original dataset announced via ProteomeXchange.

Title	De Novo Sequencing of SARS-CoV-2 and influenza monoclonal antibodies by mass spectrometry using HCD and EThcD fragmentation and Supernovo software
Description	In this study, we used electron-transfer/higher energy collision dissociation (EThcD) fragmentation to develop a mass-spectrometric (MS) method for distinguishing between Ile and Leu in Bovine Serum Albumin (BSA) and a SARS-CoV-1 anti-receptor binding domain (RBD) Spike monoclonal antibody (mAb). Supplemental activation normalized collision energy (SA-NCE) values were optimized. Combinations of multiple enzyme digestions, coupled with higher-energy collisional dissociation (HCD) and EThcD fragmentation, were used to increase coverage of all amino acid residues of the antibody. The data was searched with Supernovo™, which generated a complete de novo sequence for the test mAb and two additional SARS-CoV-2 human mAbs, giving correct sequences for the variable regions and selection between Ile and Leu residues. We then used the method on an additional set of twenty-five anti-hemagglutinin (HA) influenza antibodies of unknown sequence and determined high confidence sequences for >99% of the complementarity determining regions (CDRs). Importantly, the recombinant expression of these mAbs verified their binding and specificity to the HA influenza antigen. Unique glycosylation sites were also identified for eight antibodies. Our findings indicate this methodology results in almost complete antibody sequence coverage with >90% of residues determined with high confidence. Ile/Leu residues can be differentiated and glycosylation sites can be reliably identified.
HostingRepository	PRIDE
AnnounceDate	2022-06-03
AnnouncementXML	Submission_2022-06-03_15:41:51.300.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Peter Faull
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	complex glycosylation
Instrument	Orbitrap Fusion

Revision	Datetime	Status	ChangeLog Entry
0	2021-12-02 02:09:54	ID requested
⏵ 1	2022-06-03 15:41:51	announced
2	2023-11-14 08:54:00	announced	2023-11-14: Updated project metadata.

10.1021/ACS.JPROTEOME.1C00913;

submitter keyword: de novo sequencing, monoclonal antibodies, EThcD

Maria Person
contact affiliation	Biological Mass Spectrometry Facility, Center for Biomedical Research Support, The University of Austin at Texas
contact email	mperson@austin.utexas.edu
lab head
Peter Faull
contact affiliation	The University of Texas at Austin
contact email	peter.faull@austin.utexas.edu
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2022/06/PXD030094

PRIDE project URI

• PRIDE
  1. PXD030094
     1. Label: PRIDE project
     2. Name: De Novo Sequencing of SARS-CoV-2 and influenza monoclonal antibodies by mass spectrometry using HCD and EThcD fragmentation and Supernovo software