Carboplatin binding to a model protein in non-NaCl conditions to eliminate partial conversion to cisplatin, and the use of different criteria to choose the resolution limit

Hen egg white lysozyme (HEWL) co-crystallisation conditions of carboplatin without sodium chloride (NaCl) have been utilised to eliminate partial conversion of carboplatin to cisplatin observed previously. Tetragonal HEWL crystals were successfully obtained in 65% MPD with 0.1M citric acid buffer at pH 4.0 including DMSO. The X-ray diffraction data resolution to be used for the model refinement was reviewed using several topical criteria together. The CC1/2 criterion implemented in XDS led to data being significant to 2.0{\AA}, compared to the data only being able to be processed to 3.0{\AA} using the Bruker software package (SAINT). Then using paired protein model refinements and DPI values based on the FreeR value, the resolution limit was fine tuned to be 2.3{\AA}. Interestingly this was compared with results from the EVAL software package which gave a resolution limit of 2.2{\AA} solely using crossing 2, but 2.8{\AA} based on the Rmerge values (60%). The structural results showed that carboplatin bound to only the N{\delta} binding site of His-15 one week after crystal growth, whereas five weeks after crystal growth, two molecules of carboplatin are bound to the His-15 residue. In summary several new results have emerged: - firstly non-NaCl conditions showed a carboplatin molecule bound to His-15 of HEWL; secondly binding of one molecule of carboplatin was seen after one week of crystal growth and two molecules were bound after five weeks of crystal growth; and thirdly the use of several criteria to determine the diffraction resolution limit led to the successful use of data to higher resolution.Comment: 14 pages; submitted to Acta Cryst D Biological Crystallography reference number tz504

VP24-Karyopherin alpha binding affinities differ between Ebolavirus species, nfluencing interferon inhibition and VP24 stability

Zaire ebolavirus (EBOV), Bundibugyo ebolavirus (BDBV), and Reston ebolavirus (RESTV) belong to the same genus but exhibit different virulence properties. VP24 protein, a structural protein present in all family members, blocks interferon (IFN) signaling and likely contributes to virulence. Inhibition of IFN signaling by EBOV VP24 (eVP24) involves its interaction with the NPI-1 subfamily of karyopherin alpha (KPNA) nuclear transporters. Here, we evaluated eVP24, BDBV VP24 (bVP24), and RESTV VP24 (rVP24) interactions with three NPI-1 subfamily KPNAs (KPNA1, KPNA5, and KPNA6). Using purified proteins, we demonstrated that each VP24 binds to each of the three NPI-1 KPNAs. bVP24, however, exhibited approximately 10-fold-lower KPNA binding affinity than either eVP24 or rVP24. Cell-based assays also indicate that bVP24 exhibits decreased KPNA interaction, decreased suppression of IFN induced gene expression, and a decreased half-life in transfected cells compared to eVP24 or rVP24. Amino acid sequence alignments between bVP24 and eVP24 also identified residues within and surrounding the previously defined eVP24-KPNA5 binding interface that decrease eVP24-KPNA affinity or bVP24-KPNA affinity. VP24 mutations that lead to reduced KPNA binding affinity also decrease IFN inhibition and shorten VP24 half-lives. These data identify novel functional differences in VP24-KPNA interaction and reveal a novel impact of the VP24-KPNA interaction on VP24 stability. IMPORTANCE The interaction of Ebola virus (EBOV) VP24 protein with host karyopherin alpha (KPNA) proteins blocks type I interferon (IFN) signaling, which is a central component of the host innate immune response to viral infection. Here, we quantitatively compared the interactions of VP24 proteins from EBOV and two members of the Ebolavirus genus, Bundibugyo virus (BDBV) and Reston virus (RESTV). The data reveal lower binding affinity of the BDBV VP24 (bVP24) for KPNAs and demonstrate that the interaction with KPNA modulates inhibition of IFN signaling and VP24 stability. The effect of KPNA interaction on VP24 stability is a novel functional consequence of this virus-host interaction, and the differences identified between viral species may contribute to differences in pathogenesis

In-situ investigations of structural changes during cyclic loading by high resolution reciprocal space mapping

A major failure reason for structural materials is fatigue-related damage due to repeatedly changing mechanical loads. During cyclic loading dislocations self-organize into characteristic ordered structures, which play a decisive role for the materials lifetime. These heterogeneous dislocation structures can be identified using advanced electron microscopy and synchrotron techniques. A detailed characterization of the microstructure during cyclic loading by in-situ monitoring the internal structure within individual grains with high energy x-rays can help to understand and predict the materials behavior during cyclic deformation and to improve the material design. While monitoring macroscopic stress and strain during cyclic loading, reciprocal space maps of diffraction peaks from single grains are obtained with high resolution. High Resolution Reciprocal Space Mapping was applied successfully in-situ during cyclic deformation of macroscopic aluminium samples at the Advanced Photon Source to reveal the structural reorganization within single grains embedded in the bulk material during fatigue

Analysis of time-dependent deformation in tunnels using the Convergence-Confinement Method

During the excavation of a tunnel the accumulated wall displacement and the loading of tunnel support is the result of both the tunnel advance (round length and cycle time) and the time-dependent behaviour of the surrounding rock mass. The current approach to analyze the tunnel wall displacement increase is based on the Convergence-Confinement Method (CCM) performed with either analytical (closed form solutions) or the usage of the Longitudinal Displacement Profiles. This approach neglects the influence of time-dependency resulting in delayed deformation that may manifest even minutes or hours after excavation. Failure to consider the added displacements in the preliminary design can result in false selecting the time of installation and the type of support system causing safety issues to the working personnel, leading to cost overruns and project delivery delays. This study focuses on investigating and analyzing the total displacements around a circular tunnel in a visco-elastic medium by performing an isotropic axisymmetric finite difference modelling, proposing a new yet simplified approach that practitioners can use taking into account the effect of time

Self-stabilizing positron acceleration in a plasma column

Plasma accelerators sustain extreme field gradients, and potentially enable future compact linear colliders. Although tremendous progress has been achieved in accelerating electron beams in a plasma accelerator, positron acceleration with collider-relevant parameters is challenging. A recently proposed positron acceleration scheme relying on the wake generated by an electron drive beam in a plasma column has been shown to be able to accelerate positron witness beams with low emittance and low energy spread. However, since this scheme relies on cylindrical symmetry, it is possibly prone to transverse instabilities that could lead, ultimately, to beam break-up. In this article, we show that the witness beam itself is subject to various damping mechanisms and, therefore, this positron acceleration scheme is inherently stable towards misalignment of the drive and witness beams. This enables stable, high-quality plasma-based positron acceleration

Assessing Retinal Structure In Complete Congenital Stationary Night Blindness and Oguchi Disease

Purpose To examine retinal structure and changes in photoreceptor intensity after dark adaptation in patients with complete congenital stationary night blindness and Oguchi disease. Design Prospective, observational case series. Methods We recruited 3 patients with complete congenital stationary night blindness caused by mutations in GRM6, 2 brothers with Oguchi disease caused by mutations in GRK1, and 1 normal control. Retinal thickness was measured from optical coherence tomography images. Integrity of the rod and cone mosaic was assessed using adaptive optics scanning light ophthalmoscopy. We imaged 5 of the patients after a period of dark adaptation and examined layer reflectivity on optical coherence tomography in a patient with Oguchi disease under light- and dark-adapted conditions. Results Retinal thickness was reduced in the parafoveal region in patients with GRM6 mutations as a result of decreased thickness of the inner retinal layers. All patients had normal photoreceptor density at all locations analyzed. On removal from dark adaptation, the intensity of the rods (but not cones) in the patients with Oguchi disease gradually and significantly increased. In 1 Oguchi disease patient, the outer segment layer contrast on optical coherence tomography was 4-fold higher under dark-adapted versus light-adapted conditions. Conclusions The selective thinning of the inner retinal layers in patients with GRM6 mutations suggests either reduced bipolar or ganglion cell numbers or altered synaptic structure in the inner retina. Our finding that rods, but not cones, change intensity after dark adaptation suggests that fundus changes in Oguchi disease are the result of changes within the rods as opposed to changes at a different retinal locus

Spectral fingerprint of quantum confinement in single CsPbBr3_3 nanocrystals

Lead halide perovskite nanocrystals (NCs) are promising materials for classical and quantum lightemission applications. To gain a better understanding of their outstanding properties, a thorough understanding of the band-edge exciton emission is needed which is not reachable in ensemble and room temperature studies because of broadening effects. Here, we report on a study of the photoluminescence (PL) of single CsPbBr$_3$ NCs in the intermediate quantum confinement regime at cryogenic temperature. We reveal the size-dependence of the spectral features observed in single NCs PL: the bright-triplet exciton energy splittings, the trion and biexciton binding energies as well as the optical phonon replica spectrum. In addition, we show that the bright triplet energy splittings are consistent with a pure exchange model and that the variety of polarisation properties and PL spectra of single CsPbBr$_3$ NCs can be simply rationalised by considering the orientation of the emitting dipoles and the thermal populations of the emitting states