RT Journal Article
SR Electronic
T1 Quantitative Cross-Linking/Mass Spectrometry Reveals Subtle Protein Conformational Changes
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 055418
DO 10.1101/055418
A1 Zhuo A. Chen
A1 Lutz Fischer
A1 Salman Tahir
A1 Jimi-Carlo Bukowski-Wills
A1 Paul N. Barlow
A1 Juri Rappsilber
YR 2016
UL http://biorxiv.org/content/early/2016/05/25/055418.abstract
AB We have developed quantitative cross-linking/mass spectrometry (QCLMS) to interrogate conformational rearrangements of proteins in solution. Our workflow was tested using a structurally well-described reference system, the human complement protein C3 and its activated cleavage product C3b. We found that small local conformational changes affect the yields of cross-linking residues that are near in space while larger conformational changes affect the detectability of cross-links. Distinguishing between minor and major changes required robust analysis based on replica analysis and a label-swapping procedure. By providing workflow, code of practice and a framework for semi-automated data processing, we lay the foundation for QCLMS as a tool to monitor the domain choreography that drives binary switching in many protein-protein interaction networks.BS3Bis[sulfosuccinimidyl] suberateCLMSCross-linking/mass spectrometryFDRFalse discovery rateHCDHigher energy collision induced dissociationLC-MS/MSLiquid chromatography tandem mass spectrometryLTQLinear trap quadrupoleMS2Tandem mass spectrometryQCLMSQuantitative cross-linking/mass spectrometrySCXStrong cation exchange