A Practical Guide to iSPOT Modeling: An Integrative Structural Biology Platform
Integrative structure modeling is an emerging method for structural determination of protein-protein complexes that are challenging for conventional structural techniques. Here, we provide a practical protocol for implementing our integrated iSPOT platform by integrating three different biophysical techniques: small-angle X-ray scattering (SAXS), hydroxyl radical footprinting, and computational docking simulations. Specifically, individual techniques are described from experimental and/or computational perspectives, and complementary structural information from these different techniques are integrated for accurate characterization of the structures of large protein-protein complexes.
KeywordsiSPOT Integrative structural biology SAXS Hydroxyl radical footprinting Computational docking simulation Structural mass spectrometry Protein-protein interaction
This work was supported by the NIH (R01GM114056 and P30EB009998), the DoD (W81XWH-11-1033), and by the Ministry of Education of Singapore (2014-T2-1-065). Beamtime access was supported via the BioCAT at the Advanced Photo Source and the LiX beamline at the NSLS-II by the DoE (DE-AC02-06CH11357 and KP1605010) and by the NIH (9P41GM103622 and P41GM111244). Additional support was provided by the Ohio Supercomputer Center and the Clinical and Translational Science Collaborative of Cleveland (4UL1TR000439). The content is solely the responsibility of the authors and does not necessarily reflect the official views of the National Institute of General Medical Sciences or the National Institutes of Health.
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