Isozyme-Specific Ligands for O-acetylserine sulfhydrylase, a Novel Antibiotic Target

Conceived and designed the experiments: FS PC BC ES AM. Performed the experiments: FS RS ES PF SR. Analyzed the data: FS BC ES PF GEK PFC AM. Contributed reagents/materials/analysis tools: PC PB GC. Wrote the paper: FS GEK BC AM.The last step of cysteine biosynthesis in bacteria and plants is catalyzed by O-acetylserine sulfhydrylase. In bacteria, two isozymes, O-acetylserine sulfhydrylase-A and O-acetylserine sulfhydrylase-B, have been identified that share similar binding sites, although the respective specific functions are still debated. O-acetylserine sulfhydrylase plays a key role in the adaptation of bacteria to the host environment, in the defense mechanisms to oxidative stress and in antibiotic resistance. Because mammals synthesize cysteine from methionine and lack O-acetylserine sulfhydrylase, the enzyme is a potential target for antimicrobials. With this aim, we first identified potential inhibitors of the two isozymes via a ligand- and structure-based in silico screening of a subset of the ZINC library using FLAP. The binding affinities of the most promising candidates were measured in vitro on purified O-acetylserine sulfhydrylase-A and O-acetylserine sulfhydrylase-B from Salmonella typhimurium by a direct method that exploits the change in the cofactor fluorescence. Two molecules were identified with dissociation constants of 3.7 and 33 µM for O-acetylserine sulfhydrylase-A and O-acetylserine sulfhydrylase-B, respectively. Because GRID analysis of the two isoenzymes indicates the presence of a few common pharmacophoric features, cross binding titrations were carried out. It was found that the best binder for O-acetylserine sulfhydrylase-B exhibits a dissociation constant of 29 µM for O-acetylserine sulfhydrylase-A, thus displaying a limited selectivity, whereas the best binder for O-acetylserine sulfhydrylase-A exhibits a dissociation constant of 50 µM for O-acetylserine sulfhydrylase-B and is thus 8-fold selective towards the former isozyme. Therefore, isoform-specific and isoform-independent ligands allow to either selectively target the isozyme that predominantly supports bacteria during infection and long-term survival or to completely block bacterial cysteine biosynthesis.Yeshttp://www.plosone.org/static/editorial#pee

On the Anisotropic Mechanical Properties of Selective Laser-Melted Stainless Steel

The thorough description of the peculiarities of additively manufactured (AM) structures represents a current challenge for aspiring freeform fabrication methods, such as selective laser melting (SLM). These methods have an immense advantage in the fast fabrication (no special tooling or moulds required) of components, geometrical flexibility in their design, and efficiency when only small quantities are required. However, designs demand precise knowledge of the material properties, which in the case of additively manufactured structures are anisotropic and, under certain circumstances, inhomogeneous in nature. Furthermore, these characteristics are highly dependent on the fabrication settings. In this study, the anisotropic tensile properties of selective laser-melted stainless steel (1.4404, 316L) are investigated: the Young’s modulus ranged from 148 to 227 GPa, the ultimate tensile strength from 512 to 699 MPa, and the breaking elongation ranged, respectively, from 12% to 43%. The results were compared to related studies in order to classify the influence of the fabrication settings. Furthermore, the influence of the chosen raw material was addressed by comparing deviations on the directional dependencies reasoned from differing microstructural developments during manufacture. Stainless steel was found to possess its maximum strength at a 45° layer versus loading offset, which is precisely where AlSi10Mg was previously reported to be at its weakest

Compressive behaviour of additively manufactured AlSi10Mg

The advantages of free form fabrication methods regarding the geometrical flexibility are well known. With the full melting approach of the raw material, as utilized in selective laser melting, this freedom in design is coupled with remarkable mechanical strength. Most studies focused on the tensile characteristics; this study investigates the direction dependent compressive behaviour of selective laser melted AlSi10Mg. The obtained compressive Young's moduli exceeded both the equivalent Young's moduli for the tensile loading and the Young's modulus of the isotropic, conventionally fabricated, bulk base material, ranging as high as 82 GPa. The compressive yield strength was found to be similar to the yield point in tensile loading, with the ultimate compressive stress and strain being far superior to their tensile counterpart.No Full Tex

Additive manufacturing of cobalt-based dental alloys: analysis of microstructure and physicomechanical properties

The limitations of investment casting of cobalt-based alloys are claimed to be less problematic with significant improvements in metal additive manufacturing by selective laser melting (SLM). Despite these advantages, the metallic devices are likely to display mechanical anisotropy in relation to build orientations, which could consequently affect their performance "in vivo." In addition, there is inconclusive evidence concerning the requisite composition and postprocessing steps (e.g., heat treatment to relieve stress) that must be completed prior to using the devices. In the current paper, we evaluate the microstructure of ternary cobalt-chromium-molybdenum (Co-Cr-Mo) and cobalt-chromium-tungsten (Co-Cr-W) alloys built with direct metal printing and LaserCUSING SLM systems, respectively, at 0°, 30°, 60°, and 90° inclinations (Φ) in as-built (AB) and heat-treated (HT) conditions. The study also examines the tensile properties (Young's modulus, E; yield strength, R; elongation at failure, A; and ultimate tensile strength, R), relative density (RD), and microhardness (HV5) and macrohardness (HV20) as relevant physicomechanical properties of the alloys. Data obtained indicate improved tensile properties and HV values after a short and cost-effective heat-treatment cycle of Co-Cr-Mo alloys; however, the process did not homogenize the microstructure of the alloy. Annealing heat treatment of Co-Cr-W led to significant isotropic characteristics with increased E and A (except for Φ = 90°) in contrast to decreased R, R, and HV values, compared to the AB form. Similarly, the interlaced weld-bead structures in AB Co-Cr-W were removed during heat treatment, which led to a complete recrystallization of the microstructure. Both alloys exhibited defect-free microstructures with RD exceeding 99.5%