Explore the vast range of titles available.

Alcohol dehydrogenases (ADHs) catalyze the reversible reduction of a carbonyl group to its corresponding alcohol. ADHs are widely employed for organic synthesis due to their lack of harm to the environment, broad substrate acceptance, and high enantioselectivity. This review focuses on the impact and relevance of ADH enantioselectivities on their biotechnological application. Stereoselective ADHs are beneficial to reduce challenging ketones such as ketones owning two bulky substituents or similar-sized substituents to the carbonyl carbon. Meanwhile, in cascade reactions, non-stereoselective ADHs can be utilized for the quantitative oxidation of racemic alcohol to ketone and dynamic kinetic resolution.

Large volumes of high-throughput sequencing data have been submitted to the Sequencing Read Archive (SRA). The lack of experimental metadata associated with the data makes reuse and understanding data quality very difficult. In the case of RNA sequencing (RNA-Seq), which reveals the presence and quantity of RNA in a biological sample at any moment, it is necessary to consider that gene expression responds over a short time interval (several seconds to a few minutes) in many organisms. Therefore, to isolate RNA that accurately reflects the transcriptome at the point of harvest, raw biological samples should be processed by freezing in liquid nitrogen, immersing in RNA stabilization reagent or lysing and homogenizing in RNA lysis buffer containing guanidine thiocyanate as soon as possible. As the number of samples handled simultaneously increases, the time until the RNA is protected can increase. Here, to evaluate the effect of different lag times in RNA protection on RNA-Seq data, we harvested CHO-S cells after 3, 5, 6, and 7 days of cultivation, added RNA lysis buffer in a time course of 15, 30, 45, and 60 min after harvest, and conducted RNA-Seq. These RNA samples showed high RNA integrity number (RIN) values indicating non-degraded RNA, and sequence data from libraries prepared with these RNA samples was of high quality according to FastQC. We observed that, at the same cultivation day, global trends of gene expression were similar across the time course of addition of RNA lysis buffer; however, the expression of some genes was significantly different between the time-course samples of the same cultivation day; most of these differentially expressed genes were related to apoptosis. We conclude that the time lag between sample harvest and RNA protection influences gene expression of specific genes. It is, therefore, necessary to know not only RIN values of RNA and the quality of the sequence data but also how the experiment was performed when acquiring RNA-Seq data from the database.

Theanine, an amino acid unique to tea leaves, has been reported to exhibit stress-relieving effects. However, the stress-relieving effects of theanine (T) are greatly inhibited by caffeine (C) and epigallocatechin gallate (E), the main components of green tea, while being enhanced by arginine (A). Animal and clinical studies using matcha tea have shown that it can alleviate stress if the molar ratio of C and E against T and A (CE/TA) is less than 2. In general, the CE/TA ratio in tea leaves is reflected by the CE/TA ratio in the infused solution; however, it is not clear what infusion conditions would be expected to ensure a stress-relieving effect. In this study, to examine the stress-relieving effect of green-tea-infused solutions, the CE/TA ratio was measured under different infusion conditions. Furthermore, a study using a model solution consisting of C, E, T, and A revealed that a CE/TA ratio of at least less than 2 and a T content greater than 0.23 mM were required for stress reduction. Based on these results, we identified tea leaves and their infusion conditions that can be expected to reduce stress.

Biocatalytic approaches have gained increasing attention as sustainable alternatives to metal-catalyzed asymmetric reductions of ketones to obtain enantiopure alcohols, important intermediates for pharmaceutical synthesis. For example, enzyme-catalyzed reduction of substituted benzophenone analogs to produce chiral diaryl methanols has attracted interest, as they are the key intermediates in the synthesis of antihistamines. However, benzophenone analogs are difficult to be reduced by enzymes due to steric hindrance. Moreover, the similarities between the two groups adjacent to the carbonyl group make achieving high enantioselectivity in reduction challenging. In this study, we examined the reduction of benzophenone and its analogs by Geotrichum candidum acetophenone reductase ( Gc APRD). However, the wild type did not exhibit activity toward benzophenone due to the substrate’s bulkiness. Then, two mutants of Gc APRD (Trp288Ala and Phe56Ile/Trp288Ala) were applied to catalyze the reduction of benzophenone, resulting in high reduction yield (≥ 80%). In addition, both mutants exhibited catalytic activity toward methyl- and halogen-substituted benzophenones, especially toward 3- and 4-substituted substrates. Regarding enantioselectivity, Trp288Ala generally reduced both 3- and 4-substituted substrates to ( R )-alcohols with up to 97% ee . In contrast, Phe56Ile/Trp288Ala reduced 3-substituted substrates to ( R )-alcohols with up to 89% ee but reduced 4-substituted substrates to ( S )-alcohols with up to 92% ee . At last, the reduction mechanism was investigated using molecular docking simulations. Key points • GcAPRD mutants exhibited catalytic performance toward benzophenone analogs. • GcAPRD Phe56Ile/Trp288Ala exhibited substituent-dependent enantioselectivity. • Introducing Phe56Ile into GcAPRD Trp288Ala resulted in a clear enantiopreference. Graphical Abstract

The spider mite sub-family Tetranychinae includes many agricultural pests. The internal transcribed spacer (ITS) region of nuclear ribosomal RNA genes and the cytochrome c oxidase subunit I (COI) gene of mitochondrial DNA have been used for species identification and phylogenetic reconstruction within the sub-family Tetranychinae, although they have not always been successful. The 18S and 28S rRNA genes should be more suitable for resolving higher levels of phylogeny, such as tribes or genera of Tetranychinae because these genes evolve more slowly and are made up of conserved regions and divergent domains. Therefore, we used both the 18S (1,825-1,901 bp) and 28S (the 5' end of 646-743 bp) rRNA genes to infer phylogenetic relationships within the sub-family Tetranychinae with a focus on the tribe Tetranychini. Then, we compared the phylogenetic tree of the 18S and 28S genes with that of the mitochondrial COI gene (618 bp). As observed in previous studies, our phylogeny based on the COI gene was not resolved because of the low bootstrap values for most nodes of the tree. On the other hand, our phylogenetic tree of the 18S and 28S genes revealed several well-supported clades within the sub-family Tetranychinae. The 18S and 28S phylogenetic trees suggest that the tribes Bryobiini, Petrobiini and Eurytetranychini are monophyletic and that the tribe Tetranychini is polyphyletic. At the genus level, six genera for which more than two species were sampled appear to be monophyletic, while four genera (Oligonychus, Tetranychus, Schizotetranychus and Eotetranychus) appear to be polyphyletic. The topology presented here does not fully agree with the current morphology-based taxonomy, so that the diagnostic morphological characters of Tetranychinae need to be reconsidered.

Phylogenetic trees of spider mites were previously obtained using 18S and 28S rRNA genes. Because some of the bootstrap values were relatively low, these trees were unable to completely resolve the phylogeny. Here, we obtained RNA-Seq data for the 72 known species (73 strains) of spider mites to analyze the phylogeny of the sub-family Tetranychinae. The data were de novo assembled into a total alignment length of 790,047 bases corresponding to 264,133 amino acid residues in 652 genes. The sequence dataset was 200 times larger than the data used in the previous study. The new trees were much more robust and more clearly defined the clades of the tribes and the genera of the sub-family Tetranychinae. The tribe Tetranychini was polyphyletic because a monophyletic clade of Eurytetranychini was placed inside it. The six genera from which two or more species were sampled appeared to be monophyletic, but four genera (Schizotetranychus, Eotetranychus, Oligonychus and Tetranychus) appeared to be polyphyletic. These results strongly support the previous molecular inference of the polyphyletic tribes and genera, although the molecular phylogeny of the sub-family Tetranychinae does not fully agree with the current morphology-based taxonomy. The taxonomy of the sub-family Tetranychinae should be revised according to the molecular relationships revealed by this study.

Using PCR to distinguish closely related species can be difficult because they may have very similar genomes. Advances in bioinformatics make it possible to design PCR primers that are species-specific. In this study, we developed a bioinformatics method for extracting species-specific primer candidate sequences (i.e., unpaired primers that were specific to a single species) from RNA-Seq data sets of 19 species of spider mites (Acari, Tetranychidae). Using k -mer counting, we obtained between 257 and 48,621 species-specific unpaired primer candidates for the 19 species. We then manually obtained a second primer that was also species-specific. The primer pairs were then confirmed to work in the target species and not to work in the non-target species. Finally, species-specific primer pairs were obtained for 17 of the 19 species tested. Such species-specific primers may be used for practical species discrimination by optimizing multiplex PCR. Our primer design method is expected to be applicable to other taxa.

The development of green catalysts, specifically biocatalysts, is crucial for building a sustainable society. To enhance the versatility of biocatalysts, the immobilization of enzymes plays a vital role as it improves their recyclability and robustness. As target enzymes to immobilize, glucose dehydrogenases and carboxylases are particularly important among various kinds of enzymes due to their involvement in two significant reactions: regeneration of the reduced form of coenzyme required for various reactions, and carboxylation reactions utilizing CO2 as a substrate, respectively. In this study, we immobilized Thermoplasma acidophilum glucose dehydrogenase (TaGDH) and T. acidophilum isocitrate dehydrogenase (TaIDH) using a previously reported method involving the formation of enzyme-inorganic hybrid nanocrystals, in the course of our continuing study focusing on carboxylation catalyzed by the free form of TaGDH and TaIDH. Subsequently, we investigated the properties of the resulting immobilized enzymes. Our results indicate the successful immobilization of TaGDH and TaIDH through the formation of hybrid nanocrystals utilizing Mn2+. The immobilization process enhanced TaIDH activity, up to 211%, while TaGDH retained 71% of its original activity. Notably, the immobilized TaGDH exhibited higher activity at temperatures exceeding 87 °C than the free TaGDH. Moreover, these immobilized enzymes could be recycled. Finally, we successfully utilized the immobilized enzymes for the carboxylation of 2-ketoglutaric acid under 1 MPa CO2. In conclusion, this study represents the first immobilization of TaGDH and TaIDH using the hybrid nanocrystal forming method. Furthermore, we achieved significant activity enhancement of TaIDH through immobilization and demonstrated the recyclability of the immobilized enzymes.

Blau syndrome is a systemic autoinflammatory granulomatous disease caused by mutations in the nucleotide-binding oligomerization domain 2 ( NOD2 ) gene. NOD2 is an intracellular pathogen recognition receptor. Upon binding to muramyl dipeptide (MDP), NOD2 activates the NF-κB pathway, leading to the upregulation of proinflammatory cytokines. Clinical manifestations of Blau syndrome appear in patients before the age of four. Skin manifestations resolve spontaneously in some cases; however, joint and eye manifestations are progressive, and lead to serious complications, such as joint contracture and blindness. Currently, there is no specific curative treatment for the disease. Administration of high-dose oral steroids can improve clinical manifestations; however, treatments is difficult to maintain due to the severity of the side effects, especially in children. While several new therapies have been reported, including JAK inhibitors, anti-IL-6 and anti-IL-1 therapies, anti-TNF therapy plays a central role in the treatment of Blau syndrome. We recently performed an ex vivo study, using peripheral blood and induced pluripotent stem cells from patients. This study demonstrated that abnormal cytokine expression in macrophages from untreated patients requires IFNγ stimulation, and that anti-TNF treatment corrects the abnormalities associated with Blau syndrome, even in the presence of IFNγ. Therefore, although the molecular mechanisms by which the genetic mutations in NOD2 lead to granuloma formation remain unclear, it is possible that prior exposure to TNFα combined with IFNγ stimulation may provide the impetus for the clinical manifestations of Blau syndrome.

The evaluation of tube burring formability is a crucial task for finding a suitable material for tube-based automobile parts. The local strain at the ductile fracture site (fracture strain) should be evaluated for this purpose. Moreover, a cold-drawn steel tube has a strong anisotropic shaped microstructure and possibly causes anisotropic fracture strain behavior. Based on this background, the study evaluated the axial and hoop directional fracture strains of cold-drawn steel tubes using the small round-bar tensile specimen. The burnished surface ratio on the pierced surface was also investigated for possibility estimation of in-line formability inspection. As a result, three tubes are presented with inferior, nearly the same, and superior hoop directional fracture strains compared with the axial strains, where exceeding 40% deterioration in the hoop direction occurs by a combination of grain elongation and carbide aggregation. The scanning electron micrographs suggest that the microvoid growth and linkage percolated thorough carbides on the elongated grain boundaries. For the piercing test, the 30% fracture strain deterioration resulted in a 4% decrease in the burnished surface ratio on the pierced surface. This result suggested that the estimation of the pierced surface can detect material defects before the actual tube-burring process.