Abstract
BackgroundThe function of oligomeric proteins is inherently linked to their quaternary structure. In the absence of high-resolution data, low-resolution information in the form of spatial restraints can significantly contribute to the precision and accuracy of structural models obtained using computational approaches. To obtain such restraints, chemical cross-linking coupled with mass spectrometry (XL-MS) is commonly used. However, the use of XL-MS in the modeling of protein complexes comprised of identical subunits (homo-oligomers) is often hindered by the inherent ambiguity of intra- and inter-subunit connection assignment.ResultsWe present a comprehensive evaluation of (1) different methods for inter-residue distance calculations, and (2) different approaches for the scoring of spatial restraints. Our results show that using Solvent Accessible Surface distances (SASDs) instead of Euclidean distances (EUCs) greatly reduces the assignation ambiguity and delivers better modeling precision. Furthermore, ambiguous connections should be considered as inter-subunit only when the intra-subunit alternative exceeds the distance threshold. Modeling performance can also be improved if symmetry, characteristic for most homo-oligomers, is explicitly defined in the scoring function.ConclusionsOur findings provide guidelines for proper evaluation of chemical cross-linking-based spatial restraints in modeling homo-oligomeric protein complexes, which could facilitate structural characterization of this important group of proteins.
Highlights
The function of oligomeric proteins is inherently linked to their quaternary structure
Solvent accessible surface distances are better at distinguishing between intra- and inter-subunit connections Calculating SASDs instead of Euclidean distance (EUC) has been shown to improve the precision of modeling proteins and protein complexes, by more accurately defining which inter-residue distances agree with the experimental data [8, 18, 19]
We wanted to evaluate if calculating SASDs instead of EUCs can help distinguish between intra- and intersubunit cross-links in homo-oligomers
Summary
The function of oligomeric proteins is inherently linked to their quaternary structure. In the absence of high-resolution data, low-resolution information in the form of spatial restraints can significantly contribute to the precision and accuracy of structural models obtained using computational approaches. To obtain such restraints, chemical cross-linking coupled with mass spectrometry (XL-MS) is commonly used. Cross-linker, a bi-reactive chemical component, connects specific amino acid residues located at an appropriate distance in the 3D structure of the protein complex. The length of the cross-linker defines the maximal distance between the reactive ends of the cross-linked residues. In the distance restraint-guided docking, multiple models of complexes are generated and used to calculate inter-residue distances of experimentally identified cross-links. To confirm or reject individual models, distances calculated from these models are compared with the theoretical upper limit distances; if a model-derived distance fits the theoretically possible distance limit, the model could be considered as probable
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