The Uncommon Active Site of D-Amino Acid Transaminase from Haliscomenobacter hydrossis: Biochemical and Structural Insights into the New Enzyme

Among industrially important pyridoxal-5’-phosphate (PLP)-dependent transaminases of fold type IV D-amino acid transaminases are the least studied. However, the development of cascade enzymatic processes, including the synthesis of D-amino acids, renewed interest in their study. Here, we describe the identification, biochemical and structural characterization of a new D-amino acid transaminase from Haliscomenobacter hydrossis (Halhy). The new enzyme is strictly specific towards D-amino acids and their keto analogs; it demonstrates one of the highest rates of transamination between D-glutamate and pyruvate. We obtained the crystal structure of the Halhy in the holo form with the protonated Schiff base formed by the K143 and the PLP. Structural analysis revealed a novel set of the active site residues that differ from the key residues forming the active sites of the previously studied D-amino acids transaminases. The active site of Halhy includes three arginine residues, one of which is unique among studied transaminases. We identified critical residues for the Halhy catalytic activity and suggested functions of the arginine residues based on the comparative structural analysis, mutagenesis, and molecular modeling simulations. We suggested a strong positive charge in the O-pocket and the unshaped P-pocket as a structural code for the D-amino acid specificity among transaminases of PLP fold type IV. Characteristics of Halhy complement our knowledge of the structural basis of substrate specificity of D-amino acid transaminases and the sequence-structure-function relationships in these enzymes.     

Zur Frage der „Exoelektronen“-Emission bei Metallen

Ohne Zusammenfassung

---

     

Synthesis of tricyclic heterocycles via a tandem aryl alkylation/heck coupling sequence

A norbornene-mediated palladium-catalyzed sequence is described in which two alkyl-aryl bonds and one alkenyl-aryl bond are formed in one pot with use of microwave irradiation. A variety of symmetrical and unsymmetrical oxygen-, nitrogen-, silicon-, and sulfur-containing tricyclic heterocycles were synthesized from a Heck acceptor and an aryl iodide containing two tethered alkyl halides. This approach was further applied to the synthesis of a tricyclic mescaline analogue.     

Silica-coated La0.75Sr0.25MnO3 nanoparticles for magnetically driven DNA isolation

Magnetic La_0.75Sr_0.25MnO₃ nanoparticles possessing an approximately 20‐nm‐thick silica shell (LSMO(0.25)@SiO₂) were characterised and tested for the isolation of PCR‐ready bacterial DNA. The results presented here show that the nanoparticles do not interfere in PCR. DNA was apparently reversibly adsorbed on their silica shell from the aqueous phase system (16% PEG 6000–2 M NaCl). The method proposed was used for DNA isolation from complex food samples (dairy products and probiotic food supplements). The isolated DNA was compatible with PCR. The main advantages of the nanoparticles tested for routine use were their high colloidal stability allowing a more precise dosage and therefore high reproducibility of DNA isolation.     

Novel 4-chloro-or-4-bromoresorcinol-based bicyclic phosphonates

Abstract

The reaction of 4-chloro- and 4-bromoresorcinol with 2-ethoxyvinylphosphonic acid dichloroanhydride has been investigated for the first time. It has been determined that the product of the reaction is the mixture of structural isomers of bicyclic phosphonates at the ratio of 9:1. The structure and composition of the compounds have been elucidated from ¹H, ³¹P, and ¹³C NMR spectroscopy, mass-spectrometry (MALDI-TOF), as well as elemental and X-ray analysis.     

Method development for the determination of wood preservatives in commercially treated wood using gas chromatography-mass spectrometry

Fungicides and insecticides are commonly used preservatives to protect wood products against microbiological degradations. Currently, there is a lack of analytical methods addressing the quantitative determination of a wide range of wood preserving species in wood matrices. In this study, a reliable method was developed for the determination of a mixture of wood preserving agents with differing chemical structures (i.e., properties), including tebuconazole (TAZ), propiconazole (PAZ), 3-iodo-2-propynyl butylcarbamate (IPBC), and permethrin (PER), in pine wood. The analyte recoveries obtained by Soxhlet and multiple-stage sonication extractions were compared. While both extraction methods yielded similar results (80–100%), Soxhlet extraction was found to be less labor-intensive and thus preferred providing also lower RSDs of 1–6%. In comparison to methanol, commonly used as an extraction solvent for triazoles, acetone yielded similar extraction efficiencies for all analytes while reducing the time of sample concentration. The solid phase extraction method for triazoles was adapted to allow for a separation of IPBC and PER from the wood matrix. As opposed to previous studies, three recovery standards were employed, which enabled the correction of individual analyte losses during the sample preparation. The matrix-affected limits of detection (LODs) using gas chromatography with mass spectrometric detection were nearly the same for triazoles 0.07 and 0.21 ng g⁻¹ for PAZ and TAZ in sapwood and 0.18 and 0.21 ng g⁻¹ in heartwood, respectively. Higher LODs were observed for IPBC and PER: 3.9 and 1.7 ng g⁻¹ in sapwood, and 2.0 and 6.0 ng g⁻¹ in heartwood, respectively. The recoveries in the wood submitted to commercial sample treatment showed gradient distribution of analytes depending on the penetration of the treatment.     

Indomethacin: The Interplay between Structural Relaxation,Viscous Flow and Crystal Growth

Non-isothermal differential scanning calorimetry (DSC) was used to study the influences of particle size (d_aver) and heating rate (q⁺) on the structural relaxation, crystal growth and decomposition kinetics of amorphous indomethacin. The structural relaxation and decomposition processes exhibited d_aver-independent kinetics, with the q⁺ dependences based on the apparent activation energies of 342 and 106 kJ·mol⁻¹, respectively. The DSC-measured crystal growth kinetics played a dominant role in the nucleation throughout the total macroscopic amorphous-to-crystalline transformation: the change from the zero-order to the autocatalytic mechanism with increasing q⁺, the significant alteration of kinetics, with the storage below the glass transition temperature, and the accelerated crystallization due to mechanically induced defects. Whereas slow q⁺ led to the formation of the thermodynamically stable γ polymorph, fast q⁺ produced a significant amount of the metastable α polymorph. Mutual correlations between the macroscopic and microscopic crystal growth processes, and between the viscous flow and structural relaxation motions, were discussed based on the values of the corresponding activation energies. Notably, this approach helped us to distinguish between particular crystal growth modes in the case of the powdered indomethacin materials. Ediger’s decoupling parameter was used to quantify the relationship between the viscosity and crystal growth. The link between the cooperativity of structural domains, parameters of the Tool-Narayanaswamy-Moynihan relaxation model and microscopic crystal growth was proposed.     

Phytochemistry,Bioactivities of Metabolites,and Traditional Uses of Fagopyrum tataricum

In Tartary buckwheat (Fagopyrum tataricum), the edible parts are mainly grain and sprouts. Tartary buckwheat contains protecting substances, which make it possible for plants to survive on high altitudes and under strong natural ultraviolet radiation. The diversity and high content of phenolic substances are important for Tartary buckwheat to grow and reproduce under unfriendly environmental effects, diseases, and grazing. These substances are mainly flavonoids (rutin, quercetin, quercitrin, vitexin, catechin, epicatechin and epicatechin gallate), phenolic acids, fagopyrins, and emodin. Synthesis of protecting substances depends on genetic layout and on the environmental conditions, mainly UV radiation and temperature. Flavonoids and their glycosides are among Tartary buckwheat plants bioactive metabolites. Flavonoids are compounds of special interest due to their antioxidant properties and potential in preventing tiredness, diabetes mellitus, oxidative stress, and neurodegenerative disorders such as Parkinson’s disease. During the processing and production of food items, Tartary buckwheat metabolites are subjected to molecular transformations. The main Tartary buckwheat traditional food products are bread, groats, and sprouts.     

Time of acquisition and high spatial resolution mass spectrometry imaging

The field of mass spectrometry imaging (MSI) is constantly evolving to analyze a diverse array of biological systems. A common goal is the need to resolve cellular and subcellular heterogeneity with high spatial resolution. As the field continues to progress towards high spatial resolution, other parameters must be considered when developing a practical method. Here, we discuss the impacts of high spatial resolution on the time of acquisition and the associated implications they have on an MSI analysis (e.g., area of the region of interest). This work presents a brief tutorial serving to evaluate high spatial resolution MSI relative to time of acquisition and data file size.     

Organocatalytic Synthesis of Triflones Bearing Two Non-Adjacent Stereogenic Centers

Trifluoromethylsulfones (triflones) are useful compounds for synthesis and beyond. Yet, methods to access chiral triflones are scarce. Here, we present a mild and efficient organocatalytic method for the stereoselective synthesis of chiral triflones using α-aryl vinyl triflones, building blocks previously unexplored in asymmetric synthesis. The peptide-catalyzed reaction gives rise to a broad range of γ-triflylaldehydes with two non-adjacent stereogenic centers in high yields and stereoselectivities. A catalyst-controlled stereoselective protonation following a C−C bond formation is key to control over the absolute and relative configuration. Straightforward derivatization of the products into, e.g., disubstituted δ-sultones, γ-lactones, and pyrrolidine heterocycles highlights the synthetic versatility of the products.