Chemical-Shift Perturbations Reflect Bile Acid Binding to Norovirus Coat Protein: Recognition Comes in Different Flavors

Bile acids have been reported as important cofactors promoting human and murine norovirus (NoV) infections in cell culture. The underlying mechanisms are not resolved. Through the use of chemical shift perturbation (CSP) NMR experiments, we identified a low-affinity bile acid binding site of a human GII.4 NoV strain. Long-timescale MD simulations reveal the formation of a ligand-accessible binding pocket of flexible shape, allowing the formation of stable viral coat protein–bile acid complexes in agreement with experimental CSP data. CSP NMR experiments also show that this mode of bile acid binding has a minor influence on the binding of histo-blood group antigens and vice versa. STD NMR experiments probing the binding of bile acids to virus-like particles of seven different strains suggest that low-affinity bile acid binding is a common feature of human NoV and should therefore be important for understanding the role of bile acids as cofactors in NoV infection.     

Defining Dry Eye from a Clinical Perspective

Over the past decades, the number of patients with dry eye disease (DED) has increased dramatically. The incidence of DED is higher in Asia than in Europe and North America, suggesting the involvement of cultural or racial factors in DED etiology. Although many definitions of DED have been used, discrepancies exist between the various definitions of dry eye disease (DED) used across the globe. This article presents a clinical consensus on the definition of DED, as formulated in four meetings with global DED experts. The proposed new definition is as follows: “Dry eye is a multifactorial disease characterized by a persistently unstable and/or deficient tear film (TF) causing discomfort and/or visual impairment, accompanied by variable degrees of ocular surface epitheliopathy, inflammation and neurosensory abnormalities.” The key criteria for the diagnosis of DED are unstable TF, inflammation, ocular discomfort and visual impairment. This definition also recommends the assessment of ocular surface epitheliopathy and neurosensory abnormalities in each patient with suspected DED. It is easily applicable in clinical practice and should help practitioners diagnose DED consistently. This consensus definition of DED should also help to guide research and clinical trials that, to date, have been hampered by the lack of an established surrogate endpoint.     

Deliberately Losing Control of C−H Activation Processes in the Design of Small-Molecule-Fragment Arrays Targeting Peroxisomal Metabolism

Combined photochemical arylation, “nuisance effect” (S_NAr) reaction sequences have been employed in the design of small arrays for immediate deployment in medium-throughput X-ray protein–ligand structure determination. Reactions were deliberately allowed to run “out of control” in terms of selectivity; for example the ortho-arylation of 2-phenylpyridine gave five products resulting from mono- and bisarylations combined with S_NAr processes. As a result, a number of crystallographic hits against NUDT7, a key peroxisomal CoA ester hydrolase, have been identified.     

Preparation of Mn2+-Cu+ co-doped P2O5–ZnO–Al2O3 glasses and their red fluorescence properties

To make a Mn²⁺-doped red glass phosphor that can be excited with ultraviolet (UV) light of light-emitting diodes (LEDs), 60P₂O₅-35ZnO-5Al₂O₃-8MnO-xCu₂O glasses (x = 0-1.00) were prepared by a melt-quenching method at 1200-1400°C for 30-180 minutes in atmospheric air, and the redox of Mn and Cu as well as fluorescence properties were investigated. The Mn²⁺ ion was not reduced and oxidized in the melting, quenching, and annealing processes. The valence of Cu in the glasses changed in the order of 0, 1+, and 2+ with the increase in the amount of Cu₂O and in the melting temperature and time. In this study, a 60P₂O₅-35ZnO-5Al₂O₃-8MnO-0.10Cu₂O glass melted at 1250°C for 90 minutes, having the highest Cu⁺ concentration, showed the strongest Mn²⁺ red fluorescence under the UV light at 275 nm. This strong Mn²⁺ red fluorescence has been caused by the energy transfer from excited Cu⁺ ions to Mn²⁺ ions.     

Magnetic Resonance Signal Amplification by Reversible Exchange of Selective PyFALGEA Oligopeptide Ligands Towards Epidermal Growth Factor Receptors

The biorelevant PyFALGEA oligopeptide ligand, which is selective towards the epidermal growth factor receptor (EGFR), has been successfully employed as a substrate in magnetic resonance signal amplification by reversible exchange (SABRE) experiments. It is demonstrated that PyFALGEA and the iridium catalyst IMes form a PyFALGEA:IMes molecular complex. The interaction between PyFALGEA:IMes and H₂ results in a ternary SABRE complex. Selective 1D EXSY experiments reveal that this complex is labile, which is an essential condition for successful hyperpolarization by SABRE. Polarization transfer from parahydrogen to PyFALGEA is observed leading to significant enhancement of the ¹H NMR signals of PyFALGEA. Different iridium catalysts and peptides are inspected to discuss the influence of their molecular structures on the efficiency of hyperpolarization. It is observed that PyFALGEA oligopeptide hyperpolarization is more efficient when an iridium catalyst with a sterically less demanding NHC ligand system such as IMesBn is employed. Experiments with shorter analogues of PyFALGEA, that is, PyLGEA and PyEA, show that the bulky phenylalanine from the PyFALGEA oligopeptide causes steric hindrance in the SABRE complex, which hampers hyperpolarization with IMes. Finally, a single-scan ¹H NMR SABRE experiment of PyFALGEA with IMesBn revealed a unique pattern of NMR lines in the hydride region, which can be treated as a fingerprint of this important oligopeptide.     

Assembly of a DNA Origami Chinese Knot by Only 15% of the Staple Strands

As a giant leap in DNA self-assembly, DNA origami has exhibited an unprecedented ability to construct nanostructures with arbitrary shapes and sizes. In typical DNA origami, hundreds of short DNA staple strands fold a long, single-stranded (ss) DNA scaffold cooperatively into designed nanostructures. However, large numbers of DNA strands are expensive and would hinder applications such as pharmaceutical investigations because of the complicated components. Therefore, one challenge is how to reduce the number of staple strands needed to construct DNA origami. For a DNA origami structure, the scale-free folding pattern of the scaffold strand is determined by staple strands at the branching vertexes. Simple duplex regions help to define the size-related features of the origami geometry. In this study, we hypothesized that a scaffold strand can be correctly folded into a designed topology by using only staple strands involved in branching vertexes. After assembly, any remaining, flexible, single-stranded regions of the scaffold could be converted into rigid duplexes by DNA polymerase to achieve the designed geometric structures. To demonstrate the concept, we used only 18 staple strands (covering 15 % of the scaffold strand) to assemble a porous DNA nanostructure, which was visualized by atomic force microscopy (AFM). This study helps understanding of the role of cooperativity in origami folding, and provides a cost-effective approach for small-scale prototyping DNA origami.     

Crossing the Border: From Keto- to Imine Reduction in Short-Chain Dehydrogenases/Reductases

The family of NAD(P)H-dependent short-chain dehydrogenases/reductases (SDRs) comprises numerous biocatalysts capable of C=O or C=C reduction. The highly homologous noroxomaritidine reductase (NR) from Narcissus sp. aff. pseudonarcissus and Zt_SDR from Zephyranthes treatiae, however, are SDRs with an extended imine substrate scope. Comparison with a similar SDR from Asparagus officinalis (Ao_SDR) exhibiting keto-reducing activity, yet negligible imine-reducing capability, and mining the Short-Chain Dehydrogenase/Reductase Engineering Database indicated that NR and Zt_SDR possess a unique active-site composition among SDRs. Adapting the active site of Ao_SDR accordingly improved its imine-reducing capability. By applying the same strategy, an unrelated SDR from Methylobacterium sp. 77 (M77_SDR) with distinct keto-reducing activity was engineered into a promiscuous enzyme with imine-reducing activity, thereby confirming that the ability to reduce imines can be rationally introduced into members of the “classical” SDR enzyme family. Thus, members of the SDR family could be a promising starting point for protein approaches to generate new imine-reducing enzymes.     

Dual Inhibition of Pyruvate Dehydrogenase Complex and Respiratory Chain Complex Induces Apoptosis by a Mitochondria-Targeted Fluorescent Organic Arsenical in vitro and in vivo

Based on the potential therapeutic value in targeting mitochondria and the fluorophore tracing ability, a fluorescent mitochondria-targeted organic arsenical PDT-PAO-F16 was fabricated, which not only visualized the cellular distribution, but also exerted anti-cancer activity in vitro and in vivo via targeting pyruvate dehydrogenase complex (PDHC) and respiratory chain complexes in mitochondria. In details, PDT-PAO-F16 mainly accumulated into mitochondria within hours and suppressed the activity of PDHC resulting in the inhibition of ATP synthesis and thermogenesis disorder. Moreover, the suppression of respiratory chain complex I and IV accelerated the mitochondrial dysfunction leading to caspase family-dependent apoptosis. In vivo, the acute promyelocytic leukemia was greatly alleviated in the PDT-PAO-F16 treated group in APL mice model. Our results demonstrated the organic arsenical precursor with fluorescence imaging and target-anticancer efficacy is a promising anticancer drug.     

Deacylation of Calcium-Dependent Antibiotics from Streptomyces violaceoruber in Co-culture with Streptomyces sp. MG7-G1

When Streptomyces violaceoruber grows together with Streptomyces sp. MG7-G1, it reacts with strongly induced droplet production on its aerial mycelium. Initially the metabolite profile of droplets from S. violaceoruber in co-culture with Streptomyces sp. MG7-G1 was compared to samples from S. violaceoruber in single-culture by using high-performance liquid chromatography-mass spectrometry (HPLC-MS). Then, the exudate from agar plates of co-cultures and single cultures (after freezing and thawing) was also analysed. Several compounds were only observed when S. violaceoruber was grown in co-culture. Based on their high-resolution ESI mass spectra and their comparable retention times to the calcium-dependent antibiotics (CDAs) produced by S. violaceoruber, the new compounds were suspected to be deacylated calcium-dependent antibiotics (daCDAs), lacking the 2,3-epoxyhexanoyl residue of CDAs. This was verified by detailed analysis of the MS/MS spectra of the daCDAs in comparison to the CDAs. The major CDA compounds present in calcium ion-supplemented agar medium of co-cultures were daCDAs, thus suggesting that Streptomyces sp. MG7-G1 expresses a deacylase that degrades CDAs.