Analysis of Functional Dynamics of Modular Multidomain Proteins by SAXS and NMR.

Thompson MK, Ehlinger AC, Chazin WJ
Methods Enzymol. 2017 592: 49-76

PMID: 28668130 · DOI:10.1016/bs.mie.2017.03.017

Multiprotein machines drive virtually all primary cellular processes. Modular multidomain proteins are widely distributed within these dynamic complexes because they provide the flexibility needed to remodel structure as well as rapidly assemble and disassemble components of the machinery. Understanding the functional dynamics of modular multidomain proteins is a major challenge confronting structural biology today because their structure is not fixed in time. Small-angle X-ray scattering (SAXS) and nuclear magnetic resonance (NMR) spectroscopy have proven particularly useful for the analysis of the structural dynamics of modular multidomain proteins because they provide highly complementary information for characterizing the architectural landscape accessible to these proteins. SAXS provides a global snapshot of all architectural space sampled by a molecule in solution. Furthermore, SAXS is sensitive to conformational changes, organization and oligomeric states of protein assemblies, and the existence of flexibility between globular domains in multiprotein complexes. The power of NMR to characterize dynamics provides uniquely complementary information to the global snapshot of the architectural ensemble provided by SAXS because it can directly measure domain motion. In particular, NMR parameters can be used to define the diffusion of domains within modular multidomain proteins, connecting the amplitude of interdomain motion to the architectural ensemble derived from SAXS. Our laboratory has been studying the roles of modular multidomain proteins involved in human DNA replication using SAXS and NMR. Here, we present the procedure for acquiring and analyzing SAXS and NMR data, using DNA primase and replication protein A as examples.

© 2017 Elsevier Inc. All rights reserved.

MeSH Terms (9)

DNA Primase Humans Multiprotein Complexes Nuclear Magnetic Resonance, Biomolecular Protein Conformation Protein Domains Replication Protein A Scattering, Small Angle X-Ray Diffraction

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