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. 2010 Jan 28:11:64.
doi: 10.1186/1471-2105-11-64.

ICC-CLASS: isotopically-coded cleavable crosslinking analysis software suite

Evgeniy V Petrotchenko  1 , Christoph H Borchers
Affiliations

ICC-CLASS: isotopically-coded cleavable crosslinking analysis software suite

Evgeniy V Petrotchenko et al. BMC Bioinformatics. .

Abstract

Background: Successful application of crosslinking combined with mass spectrometry for studying proteins and protein complexes requires specifically-designed crosslinking reagents, experimental techniques, and data analysis software. Using isotopically-coded ("heavy and light") versions of the crosslinker and cleavable crosslinking reagents is analytically advantageous for mass spectrometric applications and provides a "handle" that can be used to distinguish crosslinked peptides of different types, and to increase the confidence of the identification of the crosslinks.

Results: Here, we describe a program suite designed for the analysis of mass spectrometric data obtained with isotopically-coded cleavable crosslinkers. The suite contains three programs called: DX, DXDX, and DXMSMS. DX searches the mass spectra for the presence of ion signal doublets resulting from the light and heavy isotopic forms of the isotopically-coded crosslinking reagent used. DXDX searches for possible mass matches between cleaved and uncleaved isotopically-coded crosslinks based on the established chemistry of the cleavage reaction for a given crosslinking reagent. DXMSMS assigns the crosslinks to the known protein sequences, based on the isotopically-coded and un-coded MS/MS fragmentation data of uncleaved and cleaved peptide crosslinks.

Conclusion: The combination of these three programs, which are tailored to the analytical features of the specific isotopically-coded cleavable crosslinking reagents used, represents a powerful software tool for automated high-accuracy peptide crosslink identification. See: http://www.creativemolecules.com/CM_Software.htm.

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Figures

Figure 1
Figure 1
Scheme of a typical crosslinking experiment, showing the corresponding steps of data analysis and appropriate programs. Top right, scheme for an isotopically-coded cleavable crosslinker reagent possessing optional affinity and charge groups.
Figure 2
Figure 2
Scheme of cleavage of inter-peptide (top), dead-end (middle), and intra-peptide (bottom) crosslinks. An example with eight deuterium atoms as isotopic labels is shown. Dead-end and intra-peptide crosslinks can be identified based on specific mass losses (Mclohloss and Mcliloss, respectively), due to cleavage of the crosslinker.
Figure 3
Figure 3
Examples of isotopically-coded cleavable reagents and corresponding cleavage products. X denotes hydrogen or deuterium for the light and heavy forms of the reagents, respectively.
Figure 4
Figure 4
Graphic user interfaces of the DX, DXDX, and DXMSMS programs.
Figure 5
Figure 5
Flowchart for a crosslinking experiment with ICC-CLASS data analysis.
See this image and copyright information in PMC

References

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