Optimized fragmentation regime for diazirine photo-cross-linked peptides

Simple item page
dc.contributor.authorGiese, Sven H.
dc.contributor.authorBelsom, Adam
dc.contributor.authorRappsilber, Juri
dc.date.accessioned2021-03-11T07:52:50Z
dc.date.available2021-03-11T07:52:50Z
dc.date.issued2016-07-25
dc.description.abstractCross-linking/mass spectrometry has evolved into a robust technology that reveals structural insights into proteins and protein complexes. We leverage a new tribrid instrument with improved fragmentation capacities in a systematic comparison to identify which fragmentation method would be best for the identification of cross-linked peptides. Specifically, we explored three fragmentation methods and two combinations: collision-induced dissociation (CID), beam-type CID (HCD), electron-transfer dissociation (ETD), ETciD, and EThcD. Trypsin-digested, SDA-cross-linked human serum albumin (HSA) served as a test sample, yielding over all methods and in triplicate analysis in total 2602 matched PSMs and 1390 linked residue pairs at 5% false discovery rate, as confirmed by the crystal structure. HCD wins in number of matched peptide-spectrum-matches (958 PSMs) and identified links (446). CID is most complementary, increasing the number of identified links by 13% (58 links). HCD wins together with EThcD in cross-link site calling precision, with approximately 62% of sites having adjacent backbone cleavages that unambiguously locate the link in both peptides, without assuming any cross-linker preference for amino acids. Overall quality of spectra, as judged by sequence coverage of both peptides, is best for EThcD for the majority of peptides. Sequence coverage might be of particular importance for complex samples, for which we propose a data dependent decision tree, else HCD is the method of choice. The mass spectrometric raw data has been deposited in PRIDE (PXD003737).en
dc.identifier.eissn1520-6882
dc.identifier.issn0003-2700
dc.identifier.pmid27454319
dc.identifier.urihttps://depositonce.tu-berlin.de/handle/11303/12780
dc.identifier.urihttp://dx.doi.org/10.14279/depositonce-11580
dc.language.isoenen
dc.relation.ispartof10.14279/depositonce-12031en
dc.rights.urihttps://creativecommons.org/licenses/by/4.0/en
dc.subject.ddc570 Biowissenschaften; Biologiede
dc.subject.otherproteomicsen
dc.subject.otherstatisticsen
dc.subject.othercrosslinkingen
dc.subject.otherbioinformaticsen
dc.titleOptimized fragmentation regime for diazirine photo-cross-linked peptidesen
dc.typeArticleen
dc.type.versionpublishedVersionen
dcterms.bibliographicCitation.doi10.1021/acs.analchem.6b02082en
dcterms.bibliographicCitation.issue16en
dcterms.bibliographicCitation.journaltitleAnalytical Chemistryen
dcterms.bibliographicCitation.originalpublishernameAmerican Chemical Society (ACS)en
dcterms.bibliographicCitation.originalpublisherplaceColumbus, Ohioen
dcterms.bibliographicCitation.pageend8247en
dcterms.bibliographicCitation.pagestart8239en
dcterms.bibliographicCitation.volume88en
tub.accessrights.dnbfreeen
tub.affiliationFak. 3 Prozesswissenschaften::Inst. Biotechnologie::FG Bioanalytikde
tub.affiliation.facultyFak. 3 Prozesswissenschaftende
tub.affiliation.groupFG Bioanalytikde
tub.affiliation.instituteInst. Biotechnologiede
tub.publisher.universityorinstitutionTechnische Universität Berlinen

Files

Original bundle
Now showing 1 - 2 of 2
Thumbnail Image
Name:
02_Optimize_ETD.pdf
Size:
1.83 MB
Format:
Adobe Portable Document Format
Description:
Artile
Download
Thumbnail Image
Name:
02_Optimize_ETD_Supporting.pdf
Size:
20.33 MB
Format:
Adobe Portable Document Format
Description:
Supporting Information
Download
License bundle
Now showing 1 - 1 of 1
No Thumbnail Available
Name:
license.txt
Size:
4.9 KB
Format:
Item-specific license agreed upon to submission
Description:
Download

Collections

Publications