Permeabilized Escherichia coli Whole Cells Containing Co‐Expressed Two Thermophilic Enzymes Facilitate the Synthesis of scyllo‐Inositol from myo‐Inositol

Scyllo‐inositol (SI), a stereoisomer of inositol, is regarded as a promising therapeutic agent for Alzheimer's disease. Here, an in vitro cofactor‐balance biotransformation for the production of SI from myo‐inositol (MI) by thermophilic myo‐inositol 2‐dehydrogenase (IDH) and scyllo‐inositol 2‐dehydrogenase (SIDH) is presented. These two enzymes (i.e., IDH and SIDH from Geobacillus kaustophilus) are co‐expressed in Escherichia coli BL21(DE3), and E. coli cells containing the two enzymes are permeabilized by heat treatment as whole‐cell catalysts to convert MI to SI. After condition optimizations about permeabilized temperature, reaction temperature, and initial MI concentration, about 82 g L^?1 of SI is produced from 250 g L^?1 of MI within 24 h without any cofactor supplementation. This final titer of SI produced is the highest to the authors’ limited knowledge. This study provides a promising method for the large‐scale industrial production of SI.     

Photosynthetic Reduction of Xylose to Xylitol Using Cyanobacteria

Photosynthetic generation of reducing power makes cyanobacteria an attractive host for biochemical reduction compared to cell‐free and heterotrophic systems, which require burning of additional resources for the supply of reducing equivalent. Here, using xylitol synthesis as an example, efficient uptake and reduction of xylose photoautotrophically in Synechococcus elongatus PCC7942 are demonstrated upon introduction of an effective xylose transporter from Escherichia coli (Ec‐XylE) and the NADPH‐dependent xylose reductase from Candida boidinii (Cb‐XR). Simultaneous activation of xylose uptake and matching of cofactor specificity enabled an average xylitol yield of 0.9 g g^?1 xylose and a maximum productivity of about 0.15 g L^?1 day^?1 OD^?1 with increased level of xylose supply. While long‐term cellular maintenance still appears challenging, high‐density conversion of xylose to xylitol using concentrated resting cell further pushes the titer of xylitol formation to 33 g L^?1 in six days with 85% of maximum theoretical yield. While the results show that the unknown dissipation of xylose can be minimized when coupled to a strong reaction outlet, it remains to be the major hurdle hampering the yield despite the reported inability of cyanobacteria to metabolize xylose.     

A Site‐Specific Integration Reporter System That Enables Rapid Evaluation of CRISPR/Cas9‐Mediated Genome Editing Strategies in CHO Cells

Targeted gene knockout and site‐specific integration (SSI) are powerful genome editing techniques to improve the development of industrially relevant Chinese hamster ovary (CHO) cell lines. However, past efforts to perform SSI in CHO cells are characterized by low efficiencies. Moreover, numerous strategies proposed to boost SSI efficiency in mammalian cell types have yet to be evaluated head to head or in combination to appreciably boost efficiencies in CHO. To enable systematic and rapid optimization of genome editing methods, the SSIGNAL (s ite‐s pecific i ntegration and g en ome al teration) reporter system is developed. This tool can analyze CRISPR (clustered regularly interspaced palindromic repeats)/Cas9 (CRISPR‐associated protein 9)‐mediated disruption activity alone or in conjunction with SSI efficiency. The reporter system uses green and red dual‐fluorescence signals to indicate genotype states within four days following transfection, facilitating rapid data acquisition via standard flow cytometry instrumentation. In addition to describing the design and development of the system, two of its applications are demonstrated by first comparing transfection conditions to maximize CRISPR/Cas9 activity and subsequently assessing the efficiency of several promising SSI strategies. Due to its sensitivity and versatility, the SSIGNAL reporter system may serve as a tool to advance genome editing technology.     

Plasmonic Biosensor Controlled by DNAzyme for On‐Site Genetic Detection of Pathogens

On‐site predetection of pathogens could significantly decrease of a disease outbreak or national loss in most of the countries. However, conventional detection techniques are limited in use for on‐site detection due to the necessity of specialized skill or equipment. Therefore, it is necessary to develop a new technique that can predetect pathogens in the field without special skills or equipment. Here, a DNAzyme strategy to control a plasmonic biosensor for rapid and simple visual detection of Salmonella choleraesuis is adopted. Multicomponent DNAzyme formed by target addition can cleave the linker effectively at 50 °C. Linker cleavage induces dispersion of two DNA‐immobilized gold nanoparticles and color change. Under optimized assay conditions, the target could be detected via visual discrimination sensitively and specifically. Moreover, the biosensor shows the possibility of practical use with contaminants and a 16S rRNA real target. As a result, the proposed plasmonic biosensor can visually detect S. choleraesuis without unstable enzymes, a specialized technique, or equipment. Therefore, these advantages could allow that this biosensor would be used for on‐site predetection to lower the risk of transmission of infectious diseases.     

Molecular Targets for the Testing of COVID‐19

The pandemic outbreaks of coronavirus disease 2019 (COVID‐19), caused by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), spread all over the world in a short period of time. Efficient identification of the infection by SARS‐CoV‐2 has been one of the most important tasks to facilitate all the following counter measurements in dealing with the infectious disease. In Taiwan, a COVID‐19 Open Science Platform adheres to the spirit of open science: sharing sources, data, and methods to promote progress in academic research while corroborating findings from various disciplines has established in mid‐February 2020, for collaborative research in support of the development of detection methods, therapeutics, and a vaccine for COVID‐19. Research priorities include infection control, epidemiology, clinical characterization and management, detection methods (including viral RNA detection, viral antigen detection, and serum antibody detection), therapeutics (neutralizing antibody and small molecule drugs), vaccines, and SARS‐CoV‐2 pathogenesis. In addition, research on social ethics and the law are included to take full account of the impact of the COVID‐19 virus.     

Overexpression of ER Protein Processing and Apoptosis Regulator Genes in Human Embryonic Kidney 293 Cells Improves Gene Therapy Vectors Production

Gammaretroviral and lentiviral vectors (γ‐RV and LV) are among the most used vectors in gene therapy. Currently, human embryonic kidney (HEK) 293 cells, the manufacture platform of choice for these vectors, provide low transducing particle yields, challenging their therapeutic applications and commercialization. This work studies metabolic pathways, focusing on endoplasmic reticulum (ER) protein processing and anti‐apoptotic mechanisms, influencing vector productivity in HEK 293 cell substrates. To that end, four candidate genes—protein disulfide isomerase family A member 2 gene, heat shock protein family A (Hsp70) member 5 gene, X‐box binding protein 1 gene (ER protein processing), and B‐cell lymphoma 2 protein gene (anti‐apoptotic)—are individually stably expressed in the cells. How their overexpression level influence vector yields is analyzed by establishing cell populations with incremental genomic copies of each. γ‐RV volumetric productivity increases up to 97% when overexpressing ER protein processing genes. LV volumetric production increases 53% when overexpressing the anti‐apoptotic gene. Improvements are associated with higher cell specific productivities and dependent on gene overexpression level, highlighting the importance of fine‐tuning gene expression. Overall, this work discloses gene engineering targets enabling efficient gene therapy product manufacture showing that ER protein processing and anti‐apoptotic pathways are pivotal to producer cell performance.     

Cross-scale drivers of plant trait distributions in a fragmented forest landscape

During community assembly, plant functional traits are under selective pressure from processes operating at multiple spatial scales. However, in fragmented landscapes, there is little understanding of the relative importance of local-, patch- and landscape-scale processes in shaping trait distributions. Here, we investigate cross-scale influences of landscape change on traits that dictate plant life history strategies in re-assembling plant communities in a fragmented landscape in eastern China. Using forest dynamics plots (FDPs) on 29 land-bridge islands in which all woody plants have been georeferenced and identified to species, we characterized and derived two composite measures of trait variation, representing variation across the leaf economics spectrum and plant size. We then tested for trait shifts in response to local-, patch- and landscape-scale factors, and their potential cross-scale interactions. We found substantial community-wide trait changes along local-scale gradients (i.e. forest edge to interior): more acquisitive leaf economic traits and larger sized species occurred at edges, with a significant increase in trait means and trait range. Moreover, there were significant cross-scale interaction effects of patch and landscape variables on local-scale edge effects. Altered spatial arrangement of habitat in the surrounding landscape (i.e. declining habitat amount and increasing patch density), as well as decreasing area at the patch level, exacerbated edge effects on traits distributions. We suggest that synergistic interactions of landscape- and patch-scale processes, such as dispersal limitation, on local-scale environmental filtering at edges, together shape the spatial distributions of plant life history strategies in fragmented plant communities.     

Trigonostemons G and H,dinorditerpenoid dimers with axially chiral biaryl linkage from Trigonostemon chinensis

Trigonostemons G and H, two novel dimeric dinorditerpenoids, were isolated from the stem barks of Trigonostemon chinensis. Their planar structures and relative configurations were established by extensive analysis of spectroscopic data. Trigonostemons G and H possess a homodimeric biaryl skeleton obtained from two rearranged chiral nonracemic abietane-type dinorditerpenes through an axially chiral biaryl 11,11′-linkage. Torsional scan and computation of the transition states were carried out to estimate the rotational energy barrier, and the axial chirality (aS) was determined by time-dependent density functional theory (TDDFT) electronic circular dichroism (ECD) calculations. The positive n-π* ECD transitions of the isolated carbonyl chromophore above 300 nm could be used to determine the central chirality of trigonostemon G independently by ECD calculations of the diastereomers.     

Recognition of gluconeogenic enzymes; Icl1, Fbp1, and Mdh2 by Gid4 ligase: A molecular docking study

The pro/N‐degron pathway is an evolved protein degradation pathway through the ubiquitin‐proteasome system. It is a vital pathway to attain protein homeostasis inside the liver cells with varying glucose levels. N‐terminal proline exists in more than 300 proteins in Saccharomyces cerevisiae, but only three of them are the gluconeogenic enzymes; isocitrate lyase (Icl1), fructose‐1,6‐bisphosphatase (Fbp1), and malate dehydrogenase (Mdh2). The present in silico study aims to structurally illustrate the binding of Icl1 enzyme to Gid4 ligase concerning its peers; Fbp1 and Mdh2. Based on the molecular docking scores and interactions, one can attribute the binding stability of Gid4 with degrons, to peptides of length six up to eight from the N‐terminal. Moreover, the percent change in the docking score provides a rationale for the unique Gid4‐Icl1^1‐4 interaction. The present study provides insights on the binding attitude of Gid4 ligase to degrons of different lengths, so one will consider in designing peptidomimetics to target Gid4 ligase.     

Synthesize of β‐cyclodextrin functionalized dendritic fibrous nanosilica and its application for the removal of organic dye (malachite green)

Dye removal from industrial waste water has become an important issue. The highvisibility, undesirability and recalcitrance are the significant environmental problemfor the dyes. In the present work,β‐cyclodextrin functionalized KCC‐1 (KCC‐1‐NH‐β‐CD)was synthesized and utilized to the removal of hazardous malachite green. In order to study the morphology of the synthesized nano adsorbent, Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM) were obtained from the surface of the sample. Additionally, the functionalization of KCC‐1 with β‐cyclodextrin was confirmed with Furrier Transform Infrared spectroscopy (FTIR). The textural property of KCC‐1 was verified using nitrogen adsorption/ desorption analysis (BET equation). UV‐Vis spectroscopy utilized for the investigation of malachite green by KCC‐1‐NH‐β‐CD. Specific surface area of the adsorbent was calculated to be 140 m²/g and it can be stated that the synthesized nano adsorbent has high removal efficiency. It should be noted that the adsorption capacity of the employed nano adsorbent was more than 95%, which could be attributed to high porosity of β‐cyclodextrin functionalized KCC‐1.