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Beamline B21 at the Diamond Light Source synchrotron in the UK is a small‐angle X‐ray scattering (SAXS) beamline that specializes in high‐throughput measurements via automated sample delivery systems. A system has been developed whereby a sample can be illuminated by a focused beam of light coincident with the X‐ray beam. The system is compatible with the highly automated sample delivery system at the beamline and allows a beamline user to select a light source from a broad range of wavelengths across the UV and visible spectrum and to control the timing and duration of the light pulse with respect to the X‐ray exposure of the SAXS measurement. The intensity of the light source has been characterized across the wavelength range enabling experiments where a quantitative measure of dose is important. Finally, the utility of the system is demonstrated via measurement of several light‐responsive samples. [ABSTRACT FROM AUTHOR]

Plasminogen (Plg) is the inactive form of plasmin (Plm) that exists in two major glycoforms, referred to as glycoforms I and II (GI and GII). In the circulation, Plg assumes an activation-resistant "closed" conformation via interdomain interactions and is mediated by the lysine binding site (LBS) on the kringle (KR) domains. These inter-domain interactions can be readily disrupted when Plg binds to lysine/arginine residues on protein targets or free L-lysine and analogues. This causes Plg to convert into an "open" form, which is crucial for activation by host activators. In this study, we investigated how various ligands affect the kinetics of Plg conformational change using small-angle X-ray scattering (SAXS). We began by examining the open and closed conformations of Plg using size-exclusion chromatography (SEC) coupled with SAXS. Next, we developed a high-throughput (HTP) 96-well SAXS assay to study the conformational change of Plg. This method enables us to determine the Kopen value, which is used to directly compare the effect of different ligands on Plg conformation. Based on our analysis using Plg GII, we have found that the Kopen of ε-aminocaproic acid (EACA) is approximately three times greater than that of tranexamic acid (TXA), which is widely recognized as a highly effective ligand. We demonstrated further that Plg undergoes a conformational change when it binds to the C-terminal peptides of the inhibitor α2-antiplasmin (α2AP) and receptor Plg–RKT. Our findings suggest that in addition to the C-terminal lysine, internal lysine(s) are also necessary for the formation of open Plg. Finally, we compared the conformational changes of Plg GI and GII directly and found that the closed form of GI, which has an N-linked glycosylation, is less stable. To summarize, we have successfully determined the response of Plg to various ligand/receptor peptides by directly measuring the kinetics of its conformational changes. [ABSTRACT FROM AUTHOR]

Through an expansive international effort that involved data collection on 12 small‐angle X‐ray scattering (SAXS) and four small‐angle neutron scattering (SANS) instruments, 171 SAXS and 76 SANS measurements for five proteins (ribonuclease A, lysozyme, xylanase, urate oxidase and xylose isomerase) were acquired. From these data, the solvent‐subtracted protein scattering profiles were shown to be reproducible, with the caveat that an additive constant adjustment was required to account for small errors in solvent subtraction. Further, the major features of the obtained consensus SAXS data over the q measurement range 0–1 Å−1 are consistent with theoretical prediction. The inherently lower statistical precision for SANS limited the reliably measured q‐range to <0.5 Å−1, but within the limits of experimental uncertainties the major features of the consensus SANS data were also consistent with prediction for all five proteins measured in H2O and in D2O. Thus, a foundation set of consensus SAS profiles has been obtained for benchmarking scattering‐profile prediction from atomic coordinates. Additionally, two sets of SAXS data measured at different facilities to q > 2.2 Å−1 showed good mutual agreement, affirming that this region has interpretable features for structural modelling. SAS measurements with inline size‐exclusion chromatography (SEC) proved to be generally superior for eliminating sample heterogeneity, but with unavoidable sample dilution during column elution, while batch SAS data collected at higher concentrations and for longer times provided superior statistical precision. Careful merging of data measured using inline SEC and batch modes, or low‐ and high‐concentration data from batch measurements, was successful in eliminating small amounts of aggregate or interparticle interference from the scattering while providing improved statistical precision overall for the benchmarking data set. [ABSTRACT FROM AUTHOR]

Nanofibres are an interesting phase into which amphiphilic molecules can self-assemble. Described for a large number of synthetic lipids, they were seldom reported for natural lipids like microbial amphiphiles, known as biosurfactants. In this work, we show that the palmitic acid congener of sophorolipids (SLC16:0), one of the most studied families of biosurfactants, spontaneously forms a self-assembled fibre network (SAFiN) at pH below 6 through a pH jump process. pH-resolved in situ small-angle X-ray scattering (SAXS) shows a continuous micelle-to-fibre transition, characterized by an enhanced core–shell contrast between pH 9 and pH 7 and micellar fusion into a flat membrane between pH 7 and pH 6, approximately. Below pH 6, homogeneous, infinitely long nanofibres form by peeling off the membranes. Eventually, the nanofibre network spontaneously forms a thixotropic hydrogel with fast recovery rates after applying an oscillatory strain amplitude out of the linear viscoelastic regime: after being submitted to strain amplitudes during 5 min, the hydrogel recovers about 80% and 100% of its initial elastic modulus after, respectively, 20 s and 10 min. Finally, the strength of the hydrogel depends on the medium's final pH, with an elastic modulus fivefold higher at pH 3 than at pH 6. This article is part of the theme issue 'Bio-derived and bioinspired sustainable advanced materials for emerging technologies (part 1)'. [ABSTRACT FROM AUTHOR]

Amino acid functionalized perylene bisimides (PBIs) form self‐assembled structures in solution, the nature of which depends on the local environment. Using a high‐throughput photocatalysis setup, five PBIs are studied for the hydrogen evolution reaction (HER) under a range of conditions (pH and hole scavenger concentration) across 350 experiments to explore the relationship between supramolecular structure and photocatalytic activity. Using small angle X‐ray scattering (SAXS), NMR spectroscopy and ultraviolet‐visible (UV‐vis) absorption spectroscopy, it is shown that photocatalytic activity is determined by the nature of the self‐assembled aggregate that is formed, demonstrating the potential of self‐assembly to tune activity. There is a clear correlation between the presence of charged flexible cylindrical aggregates and the occurrence of photocatalytic H2 production, with UV–vis spectroscopy indicating that the most active structure type has a distinctive form of π‐aggregation which is proposed to enable efficient charge separation across multiple PBI units. [ABSTRACT FROM AUTHOR]

B21 is a small‐angle X‐ray scattering (SAXS) beamline with a bending magnet source in the 3 GeV storage ring at the Diamond Light Source Ltd synchrotron in the UK. The beamline utilizes a double multi‐layer monochromator and a toroidal focusing optic to deliver 2 × 1012 photons per second to a 34 × 40 µm (FWHM) focal spot at the in‐vacuum Eiger 4M (Dectris) detector. A high‐performance liquid chromatography system and a liquid‐handling robot make it possible to load solution samples into a temperature‐controlled in‐vacuum sample cell with a high level of automation. Alternatively, a range of viscous or solid materials may be loaded manually using a range of custom sample cells. A default scattering vector range from 0.0026 to 0.34 Å−1 and low instrument background make B21 convenient for measuring a wide range of biological macromolecules. The beamline has run a full user programme since 2013. [ABSTRACT FROM AUTHOR]

Radiation damage is a major limitation to synchrotron small-angle X-ray scattering analysis of biomacromolecules. Flowing the sample during exposure helps to reduce the problem, but its effectiveness in the laminar-flow regime is limited by slow flow velocity at the walls of sample cells. To overcome this limitation, the coflow method was developed, where the sample flows through the centre of its cell surrounded by a flow of matched buffer. The method permits an order-of-magnitude increase of X-ray incident flux before sample damage, improves measurement statistics and maintains low sample concentration limits. The method also efficiently handles sample volumes of a few microlitres, can increase sample throughput, is intrinsically resistant to capillary fouling by sample and is suited to static samples and size-exclusion chromatography applications. The method unlocks further potential of third-generation synchrotron beamlines to facilitate new and challenging applications in solution scattering. [ABSTRACT FROM AUTHOR]

The SAXS/WAXS beamline at the Australian Synchrotron is an advanced and flexible undulator X-ray scattering beamline used for small- and wide-angle X-ray scattering analysis on a wide variety of solids, fluids and surfaces across a diverse range of research and development fields. The beamline has numerous features that minimize the intensity of the instrument background, provide automated stable optics, and allow accurate analysis of very weakly scattering samples. The geometric and intensity requirements of a three-slit collimation system are described in detail for conventional metal and single-crystal germanium slits. Straightforward ray tracing and simple linear projections describe the observed direct beam as well as parasitic background scattering geometry of the beamline at its longest camera length, providing a methodology for the design and operation of similar beamlines. As an aid to instrument design, the limit of background intensity determined by the intensity incident on single-crystal germanium guard slit edges and its q dependence was quantified at 11 keV. Details of the beamline's implementation, underlying optical concept and measured performance are given. [ABSTRACT FROM AUTHOR]

The essential metabolic enzyme biotin protein ligase (BPL) is a potential target for the development of new antibiotics required to combat drug-resistant pathogens. Staphylococcus aureus BPL ( SaBPL) is a bifunctional protein, possessing both biotin ligase and transcription repressor activities. This positions BPL as a key regulator of several important metabolic pathways. Here, we report the structural analysis of both holo- and apo-forms of SaBPL using X-ray crystallography. We also present small-angle X-ray scattering data of SaBPL in complex with its biotin-carboxyl carrier protein substrate as well as the SaBPL:DNA complex that underlies repression. This has revealed the molecular basis of ligand (biotinyl-5′-AMP) binding and conformational changes associated with catalysis and repressor function. These data provide new information to better understand the bifunctional activities of SaBPL and to inform future strategies for antibiotic discovery. [ABSTRACT FROM AUTHOR]