Crystal structure of the MYB domain of the RAD transcription factor from Antirrhinum majus.
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Amino Acid Sequence↗
Biomedical subjects
Nicolas Burton
Publications and source records attributed to Nicolas Burton.
Crystallization and preliminary X-ray analysis of the RAD protein from Antirrhinum majus.
Crystals of the RADIALIS protein from Antirrhinum majus were grown by vapour diffusion after limited proteolysis. Mass spectrometry indicated that an 8 kDa fragment had been crystallized corresponding to the predicted MYB DNA-binding domain. X-ray data collected at room temperature were consistent with tetragonal symmetry, whereas data collected at 100 K using crystals cryoprotected by supplementing the mother liquor with ethylene glycol conformed to orthorhombic symmetry. It was subsequently shown that crystals soaked in cryoprotectants that were ;osmolality-matched' to the mother liquor retained tetragonal symmetry. Using these crystals, X-ray data were collected in-house to a maximum resolution of 2 A.
Antirrhinum↗
The Arabidopsis mutant sleepy1gar2-1 protein promotes plant growth by increasing the affinity of the SCFSLY1 E3 ubiquitin ligase for DELLA protein substrates.
DELLA proteins restrain the cell proliferation and enlargement that characterizes the growth of plant organs. Gibberellin stimulates growth via 26S proteasome-dependent destruction of DELLAs, thus relieving DELLA-mediated growth restraint. Here, we show that the Arabidopsis thaliana sleepy1gar2-1 (sly1gar2-1) mutant allele encodes a mutant subunit (sly1gar2-1) of an SCF(SLY1) E3 ubiquitin ligase complex. SLY1 (the wild-type form) and sly1gar2-1 both confer substrate specificity on this complex via specific binding to the DELLA proteins. However, sly1gar2-1 interacts more strongly with the DELLA target than does SLY1. In addition, the strength of the SCFSLY1-DELLA interaction is increased by target phosphorylation. Growth-promoting DELLA destruction is dependent on SLY1 availability, on the strength of the interaction between SLY1 and the DELLA target, and on promotion of the SCFSLY1-DELLA interaction by DELLA phosphorylation.
Alkyl and Aryl Transferases↗
EMAP and EMAGE: a framework for understanding spatially organized data.
The Edinburgh MouseAtlas Project (EMAP) is a time-series of mouse-embryo volumetric models. The models provide a context-free spatial framework onto which structural interpretations and experimental data can be mapped. This enables collation, comparison, and query of complex spatial patterns with respect to each other and with respect to known or hypothesized structure. The atlas also includes a time-dependent anatomical ontology and mapping between the ontology and the spatial models in the form of delineated anatomical regions or tissues. The models provide a natural, graphical context for browsing and visualizing complex data. The Edinburgh Mouse Atlas Gene-Expression Database (EMAGE) is one of the first applications of the EMAP framework and provides a spatially mapped gene-expression database with associated tools for data mapping, submission, and query. In this article, we describe the underlying principles of the Atlas and the gene-expression database, and provide a practical introduction to the use of the EMAP and EMAGE tools, including use of new techniques for whole body gene-expression data capture and mapping.
Animals↗
Identification and structure of the anti-sigma factor-binding domain of the disulphide-stress regulated sigma factor sigma(R) from Streptomyces coelicolor.
The extracytoplasmic function (ECF) sigma factor sigma(R) is a global regulator of redox homeostasis in the antibiotic-producing bacterium Streptomyces coelicolor, with a similar role in other actinomycetes such as Mycobacterium tuberculosis. Normally maintained in an inactive state by its bound anti-sigma factor RsrA, sigma(R) dissociates in response to intracellular disulphide-stress to direct core RNA polymerase to transcribe genes, such as trxBA and trxC that encode the enzymes of the thioredoxin disulphide reductase pathway, that re-establish redox homeostasis. Little is known about where RsrA binds on sigma(R) or how it suppresses sigma(R)-dependent transcriptional activity. Using a combination of proteolysis, surface-enhanced laser desorption ionisation mass spectrometry and pull-down assays we identify an N-terminal, approximately 10kDa domain (sigma(RN)) that encompasses region 2 of sigma(R) that represents the major RsrA binding site. We show that sigma(RN) inhibits transcription by an unrelated sigma factor and that this inhibition is relieved by RsrA binding, reaffirming that region 2 is involved in binding to core RNA polymerase but also demonstrating that the likely mechanism by which RsrA inhibits sigma(R) activity is by blocking this association. We also report the 2.4A resolution crystal structure of sigma(RN) that reveals extensive structural conservation with the equivalent region of sigma(70) from Escherichia coli as well as with the cyclin-box, a domain-fold found in the eukaryotic proteins TFIIB and cyclin A. sigma(RN) has a propensity to aggregate, due to steric complementarity of oppositely charged surfaces on the domain, but this is inhibited by RsrA, an observation that suggests a possible mode of action for RsrA which we compare to other well-studied sigma factor-anti-sigma factor systems.
Bacterial Proteins↗