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Sequential Digestion with Trypsin and Elastase in Cross-Linking Mass Spectrometry.

ABSTRACT: Cross-linking mass spectrometry has become an important approach for studying protein structures and protein-protein interactions. The amino acid compositions of some protein regions impede the detection of cross-linked residues, although it would yield invaluable information for protein modeling. Here, we report on a sequential-digestion strategy with trypsin and elastase to penetrate regions with a low density of trypsin-cleavage sites. We exploited intrinsic substrate-recognition properties of elastase to specifically target larger tryptic peptides. Our application of this protocol to the TAF4-12 complex allowed us to identify cross-links in previously inaccessible regions.

SUBMITTER: Dau T

PROVIDER: S-EPMC6458965 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

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Sequential Digestion with Trypsin and Elastase in Cross-Linking Mass Spectrometry.

Dau Therese T Gupta Kapil K Berger Imre I Rappsilber Juri J

Analytical chemistry 20190313 7

Cross-linking mass spectrometry has become an important approach for studying protein structures and protein-protein interactions. The amino acid compositions of some protein regions impede the detection of cross-linked residues, although it would yield invaluable information for protein modeling. Here, we report on a sequential-digestion strategy with trypsin and elastase to penetrate regions with a low density of trypsin-cleavage sites. We exploited intrinsic substrate-recognition properties o ...[more]

PMID: 30817130

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