The use of single isomorphous replacement (SIR) has become less widespread due to difficulties in sample preparation and the identification of isomorphous native and derivative data sets. Non-isomorphism becomes even more problematic in serial experiments, because it adds natural inter-crystal non-isomorphism to heavy-atom-soaking-induced non-isomorphism. Here, a method that can successfully address these issues (and indeed can benefit from differences in heavy-atom occupancy) and additionally significantly simplifies the SIR experiment is presented.
View Article and Find Full Text PDFID23-2 is a fixed-energy (14.2 keV) microfocus beamline at the European Synchrotron Radiation Facility (ESRF) dedicated to macromolecular crystallography. The optics and sample environment have recently been redesigned and rebuilt to take full advantage of the upgrade of the ESRF to the fourth generation Extremely Brilliant Source (ESRF-EBS).
View Article and Find Full Text PDFX-ray crystallography is the major technique used to obtain high resolution information concerning the 3-dimensional structures of biological macromolecules. Until recently, a major requirement has been the availability of relatively large, well diffracting crystals, which are often challenging to obtain. However, the advent of serial crystallography and a renaissance in multi-crystal data collection methods has meant that the availability of large crystals need no longer be a limiting factor.
View Article and Find Full Text PDFActa Crystallogr D Struct Biol
February 2019
Single-wavelength anomalous diffraction (SAD) phasing from multiple crystals can be especially challenging in samples with weak anomalous signals and/or strong non-isomorphism. Here, advantage is taken of the combinatorial diversity possible in such experiments to study the relationship between merging statistics and downstream metrics of phasing signals. It is furthermore shown that a genetic algorithm (GA) can be used to optimize the grouping of data sets to enhance weak anomalous signals based on these merging statistics.
View Article and Find Full Text PDFActa Crystallogr D Struct Biol
April 2018
Specific radiation damage can be used to determine phases de novo from macromolecular crystals. This method is known as radiation-damage-induced phasing (RIP). One limitation of the method is that the dose of individual data sets must be minimized, which in turn leads to data sets with low multiplicity.
View Article and Find Full Text PDFHow the formidable diversity of forms emerges from developmental and evolutionary processes is one of the most fascinating questions in biology. The homeodomain-containing Hox proteins were recognized early on as major actors in diversifying animal body plans. The molecular mechanisms underlying how this transcription factor family controls a large array of context- and cell-specific biological functions is, however, still poorly understood.
View Article and Find Full Text PDFActa Crystallogr D Struct Biol
September 2016
Recent advances in macromolecular crystallography have made it practical to rapidly collect hundreds of sub-data sets consisting of small oscillations of incomplete data. This approach, generally referred to as serial crystallography, has many uses, including an increased effective dose per data set, the collection of data from crystals without harvesting (in situ data collection) and studies of dynamic events such as catalytic reactions. However, selecting which data sets from this type of experiment should be merged can be challenging and new methods are required.
View Article and Find Full Text PDFThe histidine kinase (HK) domain belonging to the light-oxygen-voltage histidine kinase (LOV-HK) from Brucella abortus is a member of the HWE family, for which no structural information is available, and has low sequence identity (20%) to the closest HK present in the PDB. The `off-edge' S-SAD method in macromolecular X-ray crystallography was used to solve the structure of the HK domain from LOV-HK at low resolution from crystals in a low-symmetry space group (P21) and with four copies in the asymmetric unit (∼108 kDa). Data were collected both from multiple crystals (diffraction limit varying from 2.
View Article and Find Full Text PDFThe patterning function of Hox proteins relies on assembling protein complexes with PBC proteins, which often involves a protein motif found in most Hox proteins, the so-called Hexapeptide (HX). Hox/PBC complexes likely gained functional diversity by acquiring additional modes of interaction. Here, we structurally characterize the first HX alternative interaction mode based on the paralogue-specific UbdA motif and further functionally validate structure-based predictions.
View Article and Find Full Text PDFBesides their commonly attributed role in the maintenance of low-copy number plasmids, toxin/antitoxin (TA) loci, also called 'addiction modules', have been found in chromosomes and associated to a number of biological functions such as: reduction of protein synthesis, gene regulation and retardation of cell growth under nutritional stress. The recent discovery of TA loci in obligatory intracellular species of the Rickettsia genus has prompted new research to establish whether they work as stress response elements or as addiction systems that might be toxic for the host cell. VapBC2 is a TA locus from R.
View Article and Find Full Text PDFThe Coffin-Lowry syndrome, a rare syndromic form of X-linked mental retardation, is caused by loss-of-function mutations in the hRSK2 (RPS6KA3) gene. To further investigate RSK2 (90-kDa ribosomal S6 kinase) implication in cognitive processes, a mrsk2_KO mouse has previously been generated as an animal model of Coffin-Lowry syndrome. The aim of the present study was to identify possible neurochemical dysregulation associated with the behavioral and morphological abnormalities exhibited by mrsk2_KO mice.
View Article and Find Full Text PDF