title: XLPM: efficient algorithm for the analysis of protein-protein contacts using chemical cross-linking mass spectrometry.
abstract: BACKGROUND
  Chemical cross-linking is used for protein-protein contacts mapping and for structural analysis. One of the difficulties in cross-linking studies is the analysis of mass-spectrometry data and the assignment of the site of cross-link incorporation. The difficulties are due to higher charges of fragment ions, and to the overall low-abundance of cross-link species in the background of linear peptides. Cross-linkers non-specific at one end, such as photo-inducible diazirines, may complicate the analysis further. In this report, we design and validate a novel cross-linked peptide mapping algorithm (XLPM) and compare it to StavroX, which is currently one of the best algorithms in this class.
  
  
  RESULTS
  We have designed a novel cross-link search algorithm -XLPM - and implemented it both as an online tool and as a downloadable archive of scripts. We designed a filter based on an observation that observation of a b-ion implies observation of a complimentary y-ion with high probability (b-y filter). We validated the b-y filter on the set of linear peptides from NIST library, and demonstrate that it is an effective way to find high-quality mass spectra. Next, we generated cross-linked data from an ssDNA binding protein, Rim1with a specific cross-linker disuccinimidyl suberate, and a semi-specific cross-linker NHS-Diazirine, followed by analysis of the cross-linked products by nanoLC-LTQ-Orbitrap mass spectrometry. The cross-linked data were searched by XLPM and StavroX and the performance of the two algorithms was compared. The cross-links were mapped to the X-ray structure of Rim1 tetramer. Analysis of the mixture of NHS-Diazirine cross-linked ¹⁵N and ¹⁴N-labeled Rim1 tetramers yielded ¹⁵N-labeled to ¹⁴N-labeled cross-linked peptide pairs, corresponding to C-terminus-to-N-terminus cross-linking, demonstrating interaction between different two Rim1 tetramers. Both XLPM and StavroX were successful in identification of this interaction, with XLPM leading to a better annotation of higher-charged fragments. We also put forward a new method of estimating specificity and sensitivity of identification of a cross-linked residue in the case of a non-specific cross-linker.
  
  
  CONCLUSIONS
  The novel cross-link mapping algorithm, XLPM, considerably improves the speed and accuracy of the analysis compared to other methods. The quality selection filter based on b-to-y ions ratio proved to be an effective way to select high quality cross-linked spectra.
authors.forename: Mihir
authors.initials: M
authors.lastname: Jaiswal
authors.forename: Nathaniel
authors.initials: N
authors.lastname: Crabtree
authors.forename: Michael A
authors.initials: MA
authors.lastname: Bauer
authors.forename: Roger
authors.initials: R
authors.lastname: Hall
authors.forename: Kevin D
authors.initials: KD
authors.lastname: Raney
authors.forename: Boris L
authors.initials: BL
authors.lastname: Zybailov
mesh_terms: D000465:Algorithms; D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins / Q000737:chemistry; D006801:Humans; D013058:Mass Spectrometry; D010455:Peptides / Q000737:chemistry; D025941:Protein Interaction Mapping / Q000379:methods*; D055503:Protein Multimerization; D012984:Software; D013388:Succinimides
chemical_list: D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D010455:Peptides; D013388:Succinimides; C019358:disuccinimidyl suberate