Data-independent Acquisition Improves Quantitative Cross-linking Mass Spectrometry.
Mol Cell Proteomics
; 18(4): 786-795, 2019 04.
Article
de En
| MEDLINE
| ID: mdl-30651306
ABSTRACT
Quantitative cross-linking mass spectrometry (QCLMS) reveals structural detail on altered protein states in solution. On its way to becoming a routine technology, QCLMS could benefit from data-independent acquisition (DIA), which generally enables greater reproducibility than data-dependent acquisition (DDA) and increased throughput over targeted methods. Therefore, here we introduce DIA to QCLMS by extending a widely used DIA software, Spectronaut, to also accommodate cross-link data. A mixture of seven proteins cross-linked with bis[sulfosuccinimidyl] suberate (BS3) was used to evaluate this workflow. Out of the 414 identified unique residue pairs, 292 (70%) were quantifiable across triplicates with a coefficient of variation (CV) of 10%, with manual correction of peak selection and boundaries for PSMs in the lower quartile of individual CV values. This compares favorably to DDA where we quantified cross-links across triplicates with a CV of 66%, for a single protein. We found DIA-QCLMS to be capable of detecting changing abundances of cross-linked peptides in complex mixtures, despite the ratio compression encountered when increasing sample complexity through the addition of E. coli cell lysate as matrix. In conclusion, the DIA software Spectronaut can now be used in cross-linking and DIA is indeed able to improve QCLMS.
Mots clés
Texte intégral:
1
Collection:
01-internacional
Base de données:
MEDLINE
Sujet principal:
Spectrométrie de masse
/
Réactifs réticulants
/
Analyse de données
Type d'étude:
Prognostic_studies
Limites:
Animals
/
Humans
Langue:
En
Journal:
Mol Cell Proteomics
Sujet du journal:
BIOLOGIA MOLECULAR
/
BIOQUIMICA
Année:
2019
Type de document:
Article
Pays d'affiliation:
Allemagne