PXD017316-1

PXD017316 is an original dataset announced via ProteomeXchange.

Title	Harnessing the power of electrophoresis and chromatography: Offline coupling of reverse phase liquid chromatography-capillary zone electrophoresis-tandem mass spectrometry for peptide mapping for monoclonal antibodies
Description	RPLC-MS/MS is the present workhorse for bottom-up proteomics-based characterization. However it suffers from limited peak capacity and challenges with respect to detection of small and hydrophilic peptides. CZE-MS/MS offers an orthogonal alternative to RPLC-MS/MS but has limited sensitivity due to low sample loading. In the present work, for the first time, offline coupling of an RPLC-CZE-MS/MS has been demonstrated for peptide mapping of a monoclonal antibody-based therapeutic product. The performance of this platform has been compared to the state-of-the-art RPLC-MS/MS and CZE-MS/MS approaches. Fifteen fractions were isolated from a mAb tryptic digest using RPLC, and each fraction was analyzed by CZE-ESI-MS/MS. Results indicate that not only the RPLC-CZE-MS/MS platform identify a larger number of distinct peptides (372) than the RPLC-MS/MS (219) and CZE-MS/MS (177) platforms, it also offered significantly superior sequence coverage (99.55% vs. 89.76% and 94.21% for the heavy chain and 98.6% vs. 92.06% and 95.79% for the light chain). Also, the distribution of amino acid residues with post translational modifications was 413 for RPLC-CZE-MS/MS, 150 for RPLC-MS/MS and 94 for CZE-MS/MS. In fact, the RPLC-CZE-MS/MS system performed better than the combined datasets from RPLC-MS/MS or CZE-MS/MS. Our study shows that the traditionally used one-dimensional (1D) RPLC-MS/MS and CZE-MS/MS-based platforms may not be enough for characterization of complex molecules such as monoclonal antibodies. The proposed approach should be an essential addition to the analytical toolkit for in-depth primary structural characterization of mAbs.
HostingRepository	PRIDE
AnnounceDate	2020-03-05
AnnouncementXML	Submission_2020-03-05_08:23:12.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Anurag Singh Rathore
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: 10090;
ModificationList	monohydroxylated residue; deamidated residue
Instrument	TripleTOF 5600

Revision	Datetime	Status	ChangeLog Entry
0	2020-01-29 03:49:01	ID requested
⏵ 1	2020-03-05 08:23:14	announced

Kumar R, Shah RL, Rathore AS, Harnessing the power of electrophoresis and chromatography: Offline coupling of reverse phase liquid chromatography-capillary zone electrophoresis-tandem mass spectrometry for peptide mapping for monoclonal antibodies. J Chromatogr A, 1620():460954(2020) [pubmed]

ProteomeXchange project tag: Antibodies (B/D-HPP), Human Proteome Project

submitter keyword: Biotherapeutics, peptide mapping, RPLCMSMS,CZEMSMS

Anurag Singh Rathore
contact affiliation	Professor of Chemical Enginnering Department Indian Institute of Technology Delhi
contact email	arathore@chemical.iitd.ac.in
lab head
Anurag Singh Rathore
contact affiliation	Professor Chemical Engineering Department. Indian Institute of Technology, Delhi.
contact email	arathore@chemical.iitd.ac.in
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD017316
     1. Label: PRIDE project
     2. Name: Harnessing the power of electrophoresis and chromatography: Offline coupling of reverse phase liquid chromatography-capillary zone electrophoresis-tandem mass spectrometry for peptide mapping for monoclonal antibodies