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.     

Triphenylphosphonium (TPP)-Based Antioxidants: A New Perspective on Antioxidant Design

Mitochondrial oxidative damage and dysfunction contribute to a wide range of human diseases. Considering the limitation of conventional antioxidants and that mitochondria are the main source of reactive oxygen species (ROS) which induce oxidative damage, mitochondria-targeted antioxidants which can selectively block mitochondrial oxidative damage and prevent various types of cell death have been widely developed. As a lipophilic cation, triphenylphosphonium (TPP) has been commonly used in designing mitochondria-targeted antioxidants. Conjugated with the TPP moiety, antioxidants can achieve more than 1000-fold higher mitochondrial concentration depending on cell membrane potentials and mitochondrial membrane potentials. Herein we discuss the deficiencies of conventional antioxidants and the advantages of mitochondrial targeting, and review various types of TPP-based mitochondria-targeted antioxidants. These provide theoretical and background support for the design of new anti-oxidant.     

N-Substituted Nipecotic Acids as (S)-SNAP-5114 Analogues with Modified Lipophilic Domains

Potential mGAT4 inhibitors derived from the lead substance (S)-SNAP-5114 have been synthesized and characterized for their inhibitory potency. Variations from the parent compound included the substitution of one of its aromatic 4-methoxy and 4-methoxyphenyl groups, respectively, with a more polar moiety, including a carboxylic acid, alcohol, nitrile, carboxamide, sulfonamide, aldehyde or ketone function, or amino acid partial structures. Furthermore, it was investigated how the substitution of more than one of the aromatic 4-methoxy groups affects the potency and selectivity of the resulting compounds. Among the synthesized test substances (S)-1-{2-[(4-formylphenyl)bis(4-methoxyphenyl)-methoxy]ethyl}piperidine-3-carboxylic acid, that features a carbaldehyde function in place of one of the aromatic 4-methoxy moieties of (S)-SNAP-5114, was found to have a pIC₅₀ value of 5.89±0.07, hence constituting a slightly more potent mGAT4 inhibitor than the parent substance while showing comparable subtype selectivity.     

Design,Synthesis, and Self-Assembly Studies of a Suite of Monodisperse,Facially Amphiphilic,Protein–Dendron Conjugates

The custom design of protein–dendron amphiphilic macromolecules is at the forefront of macromolecular engineering. Macromolecules with this architecture are very interesting because of their ability to self-assemble into various biomimetic nanoscopic structures. However, to date, there are no reports on this concept due to technical challenges associated with the chemical synthesis. Towards that end, herein, a new chemical methodology for the modular synthesis of a suite of monodisperse, facially amphiphilic, protein–dendron bioconjugates is reported. Benzyl ether dendrons of different generations (G1–G4) are coupled to monodisperse cetyl ethylene glycol to form macromolecular amphiphilic activity-based probes (AABPs) with a single protein reactive functionality. Micelle-assisted protein labeling technology is utilized for site-specific conjugation of macromolecular AABPs to globular proteins to make monodisperse, facially amphiphilic, protein–dendron bioconjugates. These biohybrid conjugates have the ability to self-assemble into supramolecular protein nanoassemblies. Self-assembly is primarily mediated by strong hydrophobic interactions of the benzyl ether dendron domain. The size, surface charge, and oligomeric state of protein nanoassemblies could be systematically tuned by choosing an appropriate dendron or protein of interest. This chemical method discloses a new way to custom-make monodisperse, facially amphiphilic, protein–dendron bioconjugates.     

In situ observation of synthesized nanoparticles in ultra-dilute aerosols via X-ray scattering

In-air epitaxy of nanostructures (Aerotaxy) has recently emerged as a viable route for fast, large-scale production. In this study, we use small-angle X-ray scattering to perform direct in-flight characterizations of the first step of this process, i.e., the engineered formation of Au and Pt aerosol nanoparticles by spark generation in a flow of N₂ gas. This represents a particular challenge for characterization because the particle density can be extremely low in controlled production. The particles produced are examined during production at operational pressures close to atmospheric conditions and exhibit a lognormal size distribution ranging from 5–100 nm. The Au and Pt particle production and detection are compared. We observe and characterize the nanoparticles at different stages of synthesis and extract the corresponding dominant physical properties, including the average particle diameter and sphericity, as influenced by particle sintering and the presence of aggregates. We observe highly sorted and sintered spherical Au nanoparticles at ultra-dilute concentrations (< 5 × 10⁵ particles/cm³) corresponding to a volume fraction below 3 × 10^–10, which is orders of magnitude below that of previously measured aerosols. We independently confirm an average particle radius of 25 nm via Guinier and Kratky plot analysis. Our study indicates that with high-intensity synchrotron beams and careful consideration of background removal, size and shape information can be obtained for extremely low particle concentrations with industrially relevant narrow size distributions.

     

Separation of Indium from Acid Sulfate-Containing Solutions by Ion Exchange with Impregnated Resins

Indium separation using ion exchange resins from acidic polymetallic and very diluted solutions are investigated. Since the selectivity of commercial ion exchange resins have proven to be too low for an effective separation from solutions with high content of other metals, Lewatit® TP 208 was impregnated with common extractants to enhance its properties. By resin impregnation with D2EHPA and Cyanex 272, not only the selective indium recovery was reached but also the resin capacity was increased approx. two times. The best loading and elution performance were shown by Cyanex 272-impregnated Lewatit® TP 208, increasing the indium purity in the eluate from 0.75 % to 85 %.