title: Resolving Individual Components in Protein-RNA Complexes Using Small-Angle X-ray Scattering Experiments.
abstract: Small-angle X-ray scattering (SAXS) of protein-RNA complexes has developed into an efficient and economical approach for determining low-resolution shapes of particles in solution. Here, we demonstrate a mutliphase volumetric modeling approach capable of resolving individual components within a low-resolution shape. Through three case studies, we describe the SAXS data collecting strategies, premodeling analysis, and computational methods required for deconstructing complexes into their respective components. This chapter presents an approach using the programs ScÅtter and MONSA and custom scripts for averaging and aligning of multiple independent modeling runs. The method can image small (7kDa) masses within the context of complex and is capable of visualizing ligand-induced conformational changes. Nevertheless, computational algorithms are not without error, and we describe specific considerations during SAXS data reduction and modeling to mitigate possible false positives.
authors.affiliation: Diamond Light Source Ltd., Harwell Science & Innovation Campus, Didcot, United Kingdom. Electronic address: robert.rambo@diamond.ac.uk.
authors.forename: Robert P
authors.initials: RP
authors.lastname: Rambo
mesh_terms: D000465:Algorithms; D001665:Binding Sites; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004265:DNA Helicases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008024:Ligands; D056004:Molecular Dynamics Simulation; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012322:RNA Precursors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012326:RNA Splicing; D000072260:RNA Splicing Factors; D012343:RNA, Transfer / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D053838:Scattering, Small Angle; D012984:Software; D013816:Thermodynamics; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014961:X-Ray Diffraction; D014965:X-Rays
chemical_list: D004268:DNA-Binding Proteins; D008024:Ligands; D012322:RNA Precursors; D000072260:RNA Splicing Factors; C107467:SF1 protein, human; D014157:Transcription Factors; D004247:DNA; D012343:RNA, Transfer; C412482:SMARCAL1 protein, human; D004265:DNA Helicases
keywords: Complexes; Flexibility; MONSA; Modeling; Protein; RNA; SAXS; ScÅtter; Volume-of-correlation; Volumetric