David G. Waterman

Clustering procedures for the optimal selection of data sets from multiple crystals in macromolecular crystallography.

A systematic approach to the scaling and merging of data from multiple crystals in macromolecular crystallography is introduced and explained.

Diffraction‐geometry refinement in the DIALS framework

A comprehensive description of the methods used within the DIALS framework for diffraction-geometry refinement using predicted reflection centroids is given. Examples of the advanced features of the software are provided.

New methods for indexing multi-lattice diffraction data

A new indexing method is presented which is capable of indexing multiple crystal lattices from narrow wedges of data. The efficacy of this method is demonstrated with both semi-synthetic multi-lattice data and real multi-lattice data recorded from microcrystals of ∼1 µm in size.

Conformational flexibility and molecular interactions of an archaeal homologue of the Shwachman-Bodian-Diamond syndrome protein.

BackgroundDefects in the human Shwachman-Bodian-Diamond syndrome (SBDS) protein-coding gene lead to the autosomal recessive disorder characterised by bone marrow dysfunction, exocrine pancreatic insufficiency and skeletal abnormalities. This protein is highly conserved in eukaryotes and archaea but is not found in bacteria. Although genomic and biophysical studies have suggested involvement of this protein in RNA metabolism and in ribosome biogenesis, its interacting partners remain largely unknown.ResultsWe determined the crystal structure of the SBDS orthologue from Methanothermobacter thermautotrophicus (mthSBDS). This structure shows that SBDS proteins are highly flexible, with the N-terminal FYSH domain and the C-terminal ferredoxin-like domain capable of undergoing substantial rotational adjustments with respect to the central domain. Affinity chromatography identified several proteins from the large ribosomal subunit as possible interacting partners of mthSBDS. Moreover, SELEX (Systematic Evolution of Ligands by EXponential enrichment) experiments, combined with electrophoretic mobility shift assays (EMSA) suggest that mthSBDS does not interact with RNA molecules in a sequence specific manner.ConclusionIt is suggested that functional interactions of SBDS proteins with their partners could be facilitated by rotational adjustments of the N-terminal and the C-terminal domains with respect to the central domain. Examination of the SBDS protein structure and domain movements together with its possible interaction with large ribosomal subunit proteins suggest that these proteins could participate in ribosome function.

Estimation of errors in diffraction data measured by CCD area detectors.

A simulation of a CCD area detector is presented. The methods for error estimation on data obtained from images from such detectors by two-dimensional integration are considered and improvements incorporating a realistic instrumental response are suggested.

DIALS: implementation and evaluation of a new integration package

A new X-ray diffraction data-analysis package is presented with a description of the algorithms and examples of its application to biological and chemical crystallography.

Structure and Activity of a Novel Archaeal β-CASP Protein with N-Terminal KH Domains

Summary MTH1203, a β-CASP metallo-β-lactamase family nuclease from the archaeon Methanothermobacter thermautotrophicus, was identified as a putative nuclease that might contribute to RNA processing. The crystal structure of MTH1203 reveals that, in addition to the metallo-β-lactamase nuclease and the β-CASP domains, it contains two contiguous KH domains that are unique to MTH1203 and its orthologs. RNA-binding experiments indicate that MTH1203 preferentially binds U-rich sequences with a dissociation constant in the micromolar range. In vitro nuclease activity assays demonstrated that MTH1203 is a zinc-dependent nuclease. MTH1203 is also shown to be a dimer and, significantly, this dimerization enhances the nuclease activity. Transcription termination in archaea produces mRNA transcripts with U-rich 3′ ends that could be degraded by MTH1203 considering its RNA-binding specificity. We hypothesize that this nuclease degrades mRNAs of proteins targeted for degradation and so regulates archaeal RNA turnover, possibly in concert with the exosome.

dxtbx: the diffraction experiment toolbox

A Python/C++ library for reading image data and experimental geometry for X-ray diffraction experiments from arbitrary data sources is presented.

Crystal structure of Mil (Mth680): internal duplication and similarity between the Imp4/Brix domain and the anticodon‐binding domain of class IIa aminoacyl‐tRNA synthetases

Proteins of the Imp4/Brix superfamily are involved in ribosomal RNA processing, an essential function in all cells. We report the first structure of an Imp4/Brix superfamily protein, the Mil (for Methanothermobacter thermautotrophicus Imp4‐like) protein (gene product Mth680), from the archaeon M. thermautotrophicus. The amino‐ and carboxy‐terminal halves of Mil show significant structural similarity to one another, suggesting an origin by means of an ancestral duplication. Both halves show the same fold as the anticodon‐binding domain of class IIa aminoacyl‐tRNA synthetases, with greater conservation seen in the N‐terminal half. This structural similarity, together with the charge distribution in Mil, suggests that Imp4/Brix superfamily proteins could bind single‐stranded segments of RNA along a concave surface formed by the N‐terminal half of their β‐sheet and a central α‐helix. The crystal structure of Mil is incompatible with the presence, in the Imp4/Brix domain, of a helix–turn–helix motif that was proposed to comprise the RNA‐binding moiety of the Imp4/Brix proteins.

Robust background modelling in DIALS

The application of a robust generalized linear model framework for the modelling of reflection backgrounds in X-ray diffraction images is described.