The Relationship of Precursor Cluster Concentration in a Saturated Crystallization Solution to Long-Range Order During the Transition to the Solid Phase.

A model for the transition from disordered liquid state to the solid phase has been proposed based on establishing a correlation between the concentration of precursor clusters in a saturated solution and the features of solid phase formation. The validity of the model has been verified experimentally by simultaneously studying the oligomeric structure of lysozyme protein solutions and the peculiarities of solid phase formation from these solutions. It was shown that no solid phase is formed in the absence of precursor clusters (octamers) in solution; perfect monocrystals are formed at a small concentration of octamers; mass crystallization is observed with an increasing degree of supersaturation (and concentration of octamers); further increase in octamer concentration leads to the formation of an amorphous phase.

A combined evolutionary and structural approach to disclose the primary structural determinants essential for proneurotrophins biological functions.

The neurotrophins, i.e., Nerve Growth Factor (NGF), Brain-Derived Neurotrophic Factor (BDNF), Neurotrophin 3 (NT3) and Neurotrophin 4 (NT4), are known to play a range of crucial functions in the developing and adult peripheral and central nervous systems. Initially synthesized as precursors, i.e., proneurotrophins (proNTs), that are cleaved to release C-terminal mature forms, they act through two types of receptors, the specific Trk receptors (Tropomyosin-related kinases) and the pan-neurotrophin receptor p75NTR, to initiate survival and differentiative responses. Recently, all the proNTs but proNT4 have been demonstrated to be not just inactive precursors, but signaling ligands that mediate opposing actions in fundamental aspects of the nervous system with respect to the mature counterparts through dual-receptor complexes formation with a member of the VPS10 family and p75NTR. Despite the functional relevance, the molecular determinants underpinning the interactions between the pro-domains and their receptors are still elusive probably due to their intrinsically disordered nature. Here we present an evolutionary approach coupled to an experimental study aiming to uncover the structural and dynamical basis of the biological function displayed by proNGF, proBDNF and proNT3 but missing in proNT4. A bioinformatic analysis allowed to elucidate the functional adaptability of the proNTs family in vertebrates, identifying conserved key structural features. The combined biochemical and SAXS experiments shed lights on the structure and dynamic behavior of the human proNTs in solution, giving insights on the evolutionary conserved structural motifs, essential for the multifaceted roles of proNTs in physiological as well as in pathological contexts.

Digging deeper: structural background of PEGylated fibrin gels in cell migration and lumenogenesis.

Fibrin is a well-known tool in tissue engineering, but the structure of its modifications created to improve its properties remains undiscussed despite its importance, e.g. in designing biomaterials that ensure cell migration and lumenogenesis. We sought to uncover the structural aspects of PEGylated fibrin hydrogels shown to contribute to angiogenesis. The analysis of the small-angle X-ray scattering (SAXS) data and ab initio modeling revealed that the PEGylation of fibrinogen led to the formation of oligomeric species, which are larger at a higher PEG : fibrinogen molar ratio. The improvement of optical properties was provided by the decrease in aggregates' sizes and also by retaining the bound water. Compared to the native fibrin, the structure of the 5 : 1 PEGylated fibrin gel consisted of homogenously distributed flexible fibrils with a smaller space between them. Moreover, as arginylglycylaspartic acid (RGD) sites may be partly bound to PEG-NHS or masked because of the oligomerization, the number of adhesion sites may be slightly reduced that may provide the better cell migration and formation of continuous capillary-like structures.

X-Ray Diffraction Tomography Recovery of the 3D Displacement-Field Function of the Coulomb-Type Point Defect in a Crystal.

A successive approach to the solution of the inverse problem of the X-ray diffraction tomography (XRDT) is proposed. It is based on the semi-kinematical solution of the dynamical Takagi-Taupin equations for the σ-polarized diffracted wave amplitude. Theoretically, the case of the Coulomb-type point defect in a single crystal Si(111) under the exact conditions of the symmetric Laue diffraction for a set of the tilted X-ray topography 2D-images (2D projections) is considered provided that the plane-parallel sample is rotated around the diffraction vector [[Formula: see text]20]. The iterative simulated annealing (SA) and quasi-Newton gradient descent (qNGD) algorithm codes are used for a recovery of the 3D displacement-field function of the Coulomb-type point defect. The computer recovery data of the 3D displacement-field function related to the one XRDT 2D projection are presented. It is proved that the semi-kinematical approach to the solution of the dynamical Takagi-Taupin equations is effective for recovering the 3D displacement-field function even for the one XRDT 2D projection.

ATSAS 2.8: a comprehensive data analysis suite for small-angle scattering from macromolecular solutions.

ATSAS is a comprehensive software suite for the analysis of small-angle scattering data from dilute solutions of biological macromolecules or nanoparticles. It contains applications for primary data processing and assessment, ab initio bead modelling, and model validation, as well as methods for the analysis of flexibility and mixtures. In addition, approaches are supported that utilize information from X-ray crystallography, nuclear magnetic resonance spectroscopy or atomistic homology modelling to construct hybrid models based on the scattering data. This article summarizes the progress made during the 2.5-2.8 ATSAS release series and highlights the latest developments. These include AMBIMETER, an assessment of the reconstruction ambiguity of experimental data; DATCLASS, a multiclass shape classification based on experimental data; SASRES, for estimating the resolution of ab initio model reconstructions; CHROMIXS, a convenient interface to analyse in-line size exclusion chromatography data; SHANUM, to evaluate the useful angular range in measured data; SREFLEX, to refine available high-resolution models using normal mode analysis; SUPALM for a rapid superposition of low- and high-resolution models; and SASPy, the ATSAS plugin for interactive modelling in PyMOL. All these features and other improvements are included in the ATSAS release 2.8, freely available for academic users from https://www.embl-hamburg.de/biosaxs/software.html.

Characterization of Der p 21, a new important allergen derived from the gut of house dust mites.

BACKGROUND: The house dust mite (HDM) Dermatophagoides pteronyssinus is a major allergen source eliciting allergic asthma. The aim of the study was to identify new important HDM allergens associated with allergic asthma. METHODS: A cDNA coding for a new mite allergen, designated Der p 21, was isolated using immunoglobulin E (IgE) antibodies from patients with allergic asthma out of a D. pteronyssinus expression cDNA library and expressed in Escherichia coli. RESULTS: Circular dichroism analysis of the purified allergen showed that rDer p 21 (14 726 Da) is one of the few mite allergens with an alpha-helical secondary structure. The protein exhibited high thermal stability and refolding capacity, and, as determined by small angle X-ray scattering, formed a dimer consisting of two flat triangles. rDer p 21 bound high levels of patients' IgE antibodies and showed high allergenic activity in basophil activation experiments. Rabbit anti-Der p 21 IgG antibodies inhibited mite-allergic patients' IgE binding and allowed the ultrastructural localization of the allergen in the midgut (epithelium, lumen and faeces) of D. pteronyssinus by immunogold electron microscopy. Der p 21 revealed sequence homology with group 5 mite allergens, but IgE and IgG reactivity data and cross-inhibition studies identified it as a new mite allergen. CONCLUSIONS: Der p 21 is a new important mite allergen which is liberated into the environment via faecal particles and hence may be associated with allergic asthma.

Isolation and oligomeric composition of cytochrome c nitrite reductase from the haloalkaliphilic bacterium Thioalkalivibrio nitratireducens.

A new procedure for isolation of cytochrome c nitrite reductase from the haloalkaliphilic bacterium Thioalkalivibrio nitratireducens increasing significantly the yield of the purified enzyme is presented. The enzyme is isolated from the soluble fraction of the cell extract as a hexamer, as shown by gel filtration chromatography and small angle X-ray scattering analysis. Thermostability of the hexameric form of the nitrite reductase is characterized in terms of thermoinactivation and thermodenaturation.

Structural features of the single-stranded DNA-binding protein of Epstein-Barr virus.

We report the structural features of a C-terminal deletion construct of the Epstein-Barr virus single-stranded DNA-binding protein, Balf2 (Balf2DeltaC), which like the herpes simplex virus I encoded protein, infected cell protein 8 (ICP8), binds non-sequence specifically to single-stranded DNA (ssDNA). ICP8, in the absence of ssDNA, assembles into long filamentous structures. Removal of the 60 C-terminal amino acids of ICP8 (ICP8DeltaC) prevents the formation of such filaments, whereas addition of circular ssDNA to ICP8DeltaC induces formation of "super helical" filaments. Balf2DeltaC, which we show is a zinc-binding protein, does not form these filaments under the same conditions but does bind ssDNA in a weakly cooperative manner. Further structural comparison of both proteins in solution by small-angle X-ray scattering shows proteins with similar molecular envelopes. One major difference is the tendency of Balf2DeltaC to dimerize on different surfaces to that used for oligomerization when binding to ssDNA, and this may have implications for the mechanism of replication initiation.

Structural and functional properties of mouse proNGF.

The unprocessed pro-form of the NGF (nerve growth factor), proNGF (NGF precursor, without signal peptide), has been suggested to have additional functions distinct from its role as a promoter of protein folding, i.e. apoptosis and/or neurotrophic activity. Aiming to gain insights into the specific molecular interactions that mediate proNGF biological activity and into the structural determinants stabilizing its pro-region, rm-proNGF (recombinant mouse proNGF) was expressed in Escherichia coli, refolded in vitro and characterized by physicochemical methods. X-ray solution scattering measurements (small angle X-ray scattering) revealed that rm-proNGF is dimeric in solution and appears to be anisometric when compared with the compact structure of the NGF dimer. Two structural models, a globular crab-like shape and an elongated rod-like shape, equally fit to the experimental results, pointing to an intrinsically structural disordered pro-region of NGF. The models obtained allowed the interpretation of TrkA (tropomyosin receptor kinase A) binding and activation assays in cell cultures, shedding new light on the key role of proNGF in neuronal survival and neurodegeneration.

Structural properties of Y1-xYbxNi2B2C synthesized at high pressure: EXAFS data analysis.

Local structure of Y(1-x)Yb(x)Ni2B2C series synthesized at high pressure 8 GPa has been studied using EXAFS. Measurements were performed at the Ni K-edge in temperature range 5-300 K. The results show that the Debye-Waller factor for Ni-Ni bond in the parent YNi2B2C compound is characterized by the Einstein temperature O(E) = 350 K, while a minimum value O(E) = 300 K is reached for the compound with = 0.025, which has the highest critical temperature T(c) = 12.5K of the superconductive transition. This correlates with the further suppressing of superconductivity and with the appearance of the local magnetic moments in the investigated Y(1-x)Yb(x)Ni2B2C series for x > or = 0.05 compounds. Observed changes in the local electronic and the local crystal structure of this system as a function of Yb concentration and of temperature were explained in the frame of the band filling effect.