The many faces of radiation-induced changes
ABSTRACT During diffraction experiments even cryo-cooled protein crystals can be significantly damaged due to chemical and physical changes induced by absorbed X-ray photons. The character and scale of the observed effects depend strongly on the temperature and the composition of crystals. The absorption of radiation energy results in incremental regular changes to the crystal structure, making its impact on the process of solving the structure strongly correlated with other experimental variables. An understanding of all the dependencies is still limited and does not allow for a precise prediction of the outcome of a particular diffraction experiment. Results are presented of diffraction experiments performed under different experimental conditions. The influence of temperature and crystal composition on different characteristics of radiation damage is analyzed. The observed effects are discussed in terms of their impact on data processing and phasing procedures.
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Chapter: Chapter 11.4. DENZO and SCALEPACK[Show abstract] [Hide abstract]
ABSTRACT: This chapter describes the analysis of raw diffraction data, used to produce scaled and merged diffraction amplitudes. The general description of the process is provided within the context of how it is performed in the programs DENZO and SCALEPACK. The data analysis determines many parameters of the diffraction experiment that impact the quality of the final result, providing a flexible and robust approach to problems encountered by crystallographers. Topics covered include: diffraction from a perfect crystal lattice; autoindexing; coordinate systems; experimental assumptions; prediction of the diffraction pattern; detector diagnostics; multiplicative corrections (scaling); global or post refinement; and control through a graphical interface.International Tables for Crystallography, Volume F: Crystallography of biological macromolecules, Second on-line edition edited by E. Arnold, D. M. Himmel, M. G. Rossmann, 03/2012: chapter 11.4: pages 282-295; Wiley., ISBN: 978-0-470-66078-2
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ABSTRACT: During viral RNA synthesis, Ebola virus (EBOV) nucleoprotein (NP) alternates between an RNA-template-bound form and a template-free form to provide the viral polymerase access to the RNA template. In addition, newly synthesized NP must be prevented from indiscriminately binding to noncognate RNAs. Here, we investigate the molecular bases for these critical processes. We identify an intrinsically disordered peptide derived from EBOV VP35 (NPBP, residues 20-48) that binds NP with high affinity and specificity, inhibits NP oligomerization, and releases RNA from NP-RNA complexes in vitro. The structure of the NPBP/ΔNPNTD complex, solved to 3.7 Å resolution, reveals how NPBP peptide occludes a large surface area that is important for NP-NP and NP-RNA interactions and for viral RNA synthesis. Together, our results identify a highly conserved viral interface that is important for EBOV replication and can be targeted for therapeutic development. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.Cell Reports 04/2015; DOI:10.1016/j.celrep.2015.03.034 · 7.21 Impact Factor