Sample Preparation, Data Collection, and Preliminary Data Analysis in Biomolecular Solution X‐Ray Scattering

Alexander Grishaev1

1 Laboratory of Chemical Physics, National Institutes of Health, NIDDK, Bethesda, Maryland
Publication Name:  Current Protocols in Protein Science
Unit Number:  Unit 17.14
DOI:  10.1002/0471140864.ps1714s70
Online Posting Date:  November, 2012
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Abstract

In addition to the classic methods of structural biology—X‐ray crystallography and NMR—solution X‐ray scattering (SAXS) is playing an increasingly important role in structural investigation of biological macromolecules. However, the simultaneous ease of SAXS data collection and sophistication of its data analysis tools can present challenges to the investigator. Any sample, whether pure or contaminated, whether monodisperse or polydisperse, will yield scattering data, and it is up to the user to ensure the absence of artifacts and to choose a proper structural modeling strategy. This unit discusses experimental aspects of X‐ray solution scattering, including sample preparation and data collection, as well as the steps in data processing and preliminary analysis required to ensure the absence of artifacts. The goal is to summarize everything than can go wrong with SAXS data measurement so the user can have confidence in the data before undertaking structural modeling. Curr. Protoc. Protein Sci. 70:17.14.1‐17.14.18. © 2012 by John Wiley & Sons, Inc.

Keywords: solution X‐ray scattering; SAXS; WAXS; sample preparation; aggregation; radiation damage; protein and RNA structure; structural biology

     
 
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Table of Contents

  • Introduction
  • Preparation of the Matching Buffer for SAXS Measurements
  • Buffer Composition Guidelines for SAXS Samples
  • Final Steps in SAXS Sample Preparation
  • SAXS Data Measurement: Laboratory‐Based Instruments Versus Synchrotron Beam Lines
  • SAXS Data Measurement on a Laboratory‐Based Instrument
  • Synchrotron SAXS Data Measurement: q‐Range Selection(s)
  • Measurements of the Synchrotron SAXS Data: Preliminary Steps
  • Summary
  • Acknowledgements
  • Literature Cited
  • Figures
     
 
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Materials

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Figures

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Literature Cited

Literature Cited
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