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Mass spectrometry identifiable cross-linking strategy for studying protein-protein interactions.
- Source :
-
Analytical chemistry [Anal Chem] 2005 Jan 01; Vol. 77 (1), pp. 311-8. - Publication Year :
- 2005
-
Abstract
- A new mass spectrometry identifiable cross-linking strategy has been developed to study protein-protein interactions. The new cross-linker was designed to have two low-energy MS/MS-cleavable bonds in the spacer chain to provide three primary benefits: First, a reporter tag can be released from cross-link due to cleavage of the two labile bonds in the spacer chain. Second, a relatively simple MS/MS spectrum can be generated owing to favorable cleavage of labile bonds. And finally, the cross-linked peptide chains are dissociated from each other, and each then can be fragmented separately to get sequence information. Therefore, this novel type of cross-linker was named protein interaction reporter (PIR). To this end, two RINK groups were utilized to make our first-generation cross-linker using solid-phase peptide synthesis chemistry. The RINK group contains a bond more labile than peptide bonds during low-energy activation. The new cross-linker was applied to cross-link ribonuclease S (RNase S), a noncovalent complex of S-peptide and S-protein. The results demonstrated that the new cross-linker effectively reacted with RNase S to generate various types of cross-linked products. More importantly, the cross-linked peptides successfully released reporter ions during selective MS/MS conditions, and the dissociated peptide chains remained intact during MS(2), thus enabling MS(3) to be performed subsequently. In addition, dead-end, intra-, and inter-cross-linked peptides can be distinguished by analyzing MS/MS spectra.
Details
- Language :
- English
- ISSN :
- 0003-2700
- Volume :
- 77
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Analytical chemistry
- Publication Type :
- Academic Journal
- Accession number :
- 15623310
- Full Text :
- https://doi.org/10.1021/ac0488762