CCL28 promotes progression of hepatocellular carcinoma through PDGFD-regulated MMP9 and VEGFA pathways.

Hepatocellular carcinoma (HCC) remains a major global health concern with limited therapeutic options and poor prognosis. Chemokines have emerged as critical regulators in the progression and metastasis of HCC. This study aims to investigate the mechanisms involved in CCL28-promoted progression of HCC and provide novel therapeutic targets for HCC treatment. Relationship between CCL28 expression and HCC progression were investigated by bioinformatic analysis and immunohistochemical staining assays. CCK-8, Transwell, and colony formation assay were conducted to explore the impact of CCL28 on the growth, migration and invasion of HCC cells. Quantitative real-time PCR and western blotting assays were performed to learn potential molecular mechanisms underlying the transformation of HCC driven by CCL28. The results showed that there was a direct link between increased CCL28 levels and the advancement of HCC, leading to a worse outcome. CCL28 significantly augmented malignant transformation of HCC cells, containing proliferation, migration, invasion, and clonogenicity, via activation of PDGFD-regulated MMP9 and VEGFA pathways. CCL28 emerges as a pivotal contributor to HCC tumorigenesis, propelling HCC development through the PDGFD signaling pathway. Our findings unveil potential therapeutic targets for HCC treatment.

The association between different impact exercises and osteoporosis: an analysis of data from the Taiwan biobank.

Osteoporosis is a prevalent condition marked by reduced bone density and an elevated risk of fractures, especially among postmenopausal women. Exercise plays a crucial role in preventing and managing osteoporosis, with weight-bearing and impact exercises being particularly effective in enhancing bone density and mitigating disease risk. This study investigated the relationship between various types of impact exercises and osteoporosis using data from the Taiwan Biobank (TWB). The study sample comprised 5,123 individuals without osteoporosis and 1,770 individuals with the condition. Student's t-test and logistic regression analyses were utilized to assess the associations between exercise types and osteoporosis risk. Results indicated that high-impact exercise significantly reduced the likelihood of developing osteoporosis compared to no exercise (odds ratio; OR = 0.573, 95% CI: 0.406-0.810, P = 0.002). Conversely, low-impact exercises did not show a significant overall association with osteoporosis (OR = 1.160, 95% CI: 0.932-1.445, P = 0.184). Stratified analysis by sex revealed that high-impact exercise was protective against osteoporosis in men (OR = 0.391, 95% CI: 0.202-0.755, P = 0.005), but not significantly so in women (OR = 0.671, 95% CI: 0.438-1.027, P = 0.066). These findings suggest that high-impact exercise is associated with a reduced risk of osteoporosis, particularly among Taiwanese men aged 30 to 70.

Improving the inhibitory resistance of xylanase FgXyn11C from Fusarium graminearum to SyXIP-I by site-directed mutagenesis.

The aim of this study was to improve the inhibitory resistance of xylanase FgXyn11C from Fusarium graminearum to XIP in cereal flour. Site saturation mutagenesis was performed using computer-aided redesign. Firstly, based on multiple primary structure alignments, the amino acid residues in the active site architecture were identified, and specific residue T144 in the thumb region of FgXyn11C was selected for site-saturation mutagenesis. After screening, FgXyn11CT144F was selected as the best mutant, as it displayed the highest enzymatic activity and resistance simultaneously compared to other mutants. The specific activity of FgXyn11CT144F was 208.8 U/mg and it exhibited complete resistance to SyXIP-I. Compared with the wild-type, FgXyn11CT144F displayed similar activity and the most resistant against SyXIP-I. The optimal temperature and pH of the wild-type and purified FgXyn11CT144F were similar at pH 5.0 and 30 °C. Our findings provided preliminary insight into how the specific residue at position 144 in the thumb region of FgXyn11C influenced the enzymatic properties and interacted with SyXIP-I. The inhibition sensitivity of FgXyn11C was reduced through directed evolution, leading to creation of the mutant enzyme FgXyn11CT144F. The FgXyn11CT144F resistance to SyXIP-I has potential application and can also provide references for engineering other resistant xylanases of the GHF11.

Cloning and characterization of a novel mesophilic xylanase gene Fgxyn3 from Fusarium graminearum Z-1.

In order to search for high specific activity and the resistant xylanases to XIP-I and provide more alternative xylanases for industrial production, a strain of Fusarium graminearum from Triticum aestivum grains infected with filamentous fungus produced xylanases was isolated and identified. Three xylanase genes from Fusarium graminearum Z-1 were cloned and successfully expressed in E. coli and P. pastoris, respectively. The specific activities of Fgxyn1, EFgxyn2 and EFgxyn3 for birchwood xylan were 38.79, 0.85 and 243.83 U/mg in E. coli, and 40.11, 0 and 910.37 U/mg in P. pastoris, respectively. EFgxyn3 and PFgxyn3 had the similar optimum pH at 6.0 and pH stability at 5.0-9.0. However, they had different optimum temperature and thermal stability, with 30 °C for EFgxyn3 and 40 °C for PFgxyn3, and 4-35 °C for EFgxyn3 and 4-40 °C for PFgxyn3, respectively. The substrate spectrum and the kinetic parameters showed that the two xylanases also exhibited the highest xylanase activity and catalytic efficiency (kcat/km) toward birchwood xylan, with 243.83 U/mg and 61.44 mL/mg/s for EFgxyn3 and 910.37 U/mg and 910.37 mL/mg/s for PFgxyn3, respectively. This study provided a novel mesophilic xylanase with high specific activity and catalytic efficiency, thus making it a promising candidate for extensive applications in animal feed and food industry. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-024-03973-0.

Expression and characterization of ß-1,3-1,4-glucanase of Aspergillus usamii in Escherichia coli and its application in sourdough bread making.

A ß-1,3-1,4-glucanase gene (Auglu12A) from Aspergillus usamii was successfully expressed in Escherichia coli BL21(DE3). The recombinant enzyme, reAuglu12A was efficiently purified using the one-step nickel-nitrilotriacetic acid affinity chromatography. The specific activity of reAuglu12A was 694.8 U/mg, with an optimal temperature of 55°C and pH of 5.0. The reAuglu12A exhibited stability at temperatures up to 60°C and within the pH range of 4.0-5.5. The reAuglu12A hydrolytic activity was increased in the presence of metal ions, especially K+ and Na+ , whereas it exhibited a Km and Vmax of 8.35 mg/mL and 1254.02 µmol/min/mg, respectively, toward barley ß-glucan at pH 5.0 and 55°C. The addition of reAuglu12A significantly increased the specific volume (p < 0.05) and reduced crumb firmness and chewiness (p < 0.05) of wheat-barley sourdough bread during a 7-day storage period compared to the control. Overall, the quality of wheat-barley sourdough bread was improved after incorporation of reAuglu12A (especially at 3000 U/300 g). These changes were attributed to the synergistic effect of acidification by sourdough and its metabolites which provided a conducive environment for the optimal action of reAuglu12A in the degradation of ß-glucans of barley flour in sourdough. This stabilized the dough structure, thereby enhancing the quality, texture, and shelf life of the bread. These findings suggest that reAuglu12A holds promise as a candidate for ß-glucanase application in the baking industry.

Directed Modification of a GHF11 Thermostable Xylanase AusM for Enhancing Inhibitory Resistance towards SyXIP-I and Application of AusMPKK in Bread Making.

To reduce the inhibition sensitivity of a thermoresistant xylanase AusM to xylanase inhibitor protein (XIP)-type in wheat flour, the site-directed mutagenesis was conducted based on the computer-aided redesign. First, fourteen single-site variants and one three-amino acid replacement variant in the thumb region of an AusM-encoding gene (AusM) were constructed and expressed in E. coli BL21(DE3), respectively, as predicted theoretically. At a molar ratio of 100:1 between SyXIP-I/xylanase, the majority of mutants were nearly completely inactivated by the inhibitor SyXIP-I, whereas AusMN127A retained 62.7% of its initial activity and AusMPKK retained 100% of its initial activity. The optimal temperature of the best mutant AusMPKK was 60 °C, as opposed to 60-65 °C for AusM, while it exhibited improved thermostability, retaining approximately 60% of its residual activity after heating at 80 °C for 60 min. Furthermore, AusMPKK at a dosage of 1000 U/kg was more effective than AusM at 4000 U/kg in increasing specific bread loaf volume and reducing hardness during bread production and storage. Directed evolution of AusM significantly reduces inhibition sensitivity, and the mutant enzyme AusMPKK is conducive to improving bread quality and extending its shelf life.

Origin of Limiting and Overlimiting Currents in Bipolar Membranes.

Bipolar membranes (BPMs), a special class of ion exchange membranes with the unique ability to electrochemically induce either water dissociation or recombination, are of growing interest for environmental applications including eliminating chemical dosage for pH adjustment, resource recovery, valorization of brines, and carbon capture. However, ion transport within BPMs, and particularly at its junction, has remained poorly understood. This work aims to theoretically and experimentally investigate ion transport in BPMs under both reverse and forward bias operation modes, taking into account the production or recombination of H+ and OH-, as well as the transport of salt ions (e.g., Na+, Cl-) inside the membrane. We adopt a model based on the Nernst-Planck theory, that requires only three input parameters─membrane thickness, its charge density, and pK of proton adsorption─to predict the concentration profiles of four ions (H+, OH-, Na+, and Cl-) inside the membrane and the resulting current-voltage curve. The model can predict most of the experimental results measured with a commercial BPM, including the observation of limiting and overlimiting currents, which emerge due to particular concentration profiles that develop inside the BPM. This work provides new insights into the physical phenomena in BPMs and helps identify optimal operating conditions for future environmental applications.

An Integrated Strategy for Investigating Antioxidants from Ribes himalense Royle ex Decne and Their Potential Target Proteins.

Natural products have been used extensively around the world for many years as therapeutic, prophylactic, and health-promotive agents. Ribes himalense Royle ex Decne, a plant used in traditional Tibetan medicine, has been demonstrated to have significant antioxidant and anti-inflammatory properties. However, the material basis of its medicinal effects has not been sufficiently explored. In this study, we established an integrated strategy by online HPLC-1,1-diphenyl-2-picrylhydrazyl, medium-pressure liquid chromatography, and HPLC to achieve online detection and separation of antioxidants in Ribes himalense extracts. Finally, four antioxidants with quercetin as the parent nucleus were obtained, namely, Quercetin-3-O-ß-D-glucopyranoside-7-O-α-L-rhamnopyranoside, Quercetin-3-O-ß-D-xylopyranosyl(1-2)-ß-D-glucopyranoside, Quercetin-3-O-ß-D-glucopyranoside, and Quercetin-3-O-ß-D-galactoside. Until now, the four antioxidants in Ribes himalense have not been reported in other literatures. Meanwhile, the free-radical-scavenging ability of them was evaluated by DPPH assay, and potential antioxidant target proteins were explored using molecular docking. In conclusion, this research provides insights into the active compounds in Ribes himalense which will facilitate the advancement of deeper studies on it. Moreover, such an integrated chromatographic strategy could be a strong driver for more efficient and scientific use of other natural products in the food and pharmaceutical industries.

Identification of S100A9 as a Potential Inflammation-Related Biomarker for Radiation-Induced Lung Injury.

Radiation-induced lung injury (RILI), a potentially fatal and dose-limiting complication of radiotherapy for thoracic tumors, is divided into early reversible pneumonitis and irreversible advanced-stage fibrosis. Early detection and intervention contribute to improving clinical outcomes of patients. However, there is still a lack of reliable biomarkers for early prediction and clinical diagnosis of RILI. Given the central role of inflammation in the initiation and progression of RILI, we explored specific inflammation-related biomarkers during the development of RILI in this study. Two expression profiles from the Gene Expression Omnibus (GEO) database were downloaded, in which 75 differentially expressed genes (DEGs) were screened out. Combining Gene Oncology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and protein-protein interaction (PPI) network analysis, we identified four inflammation-related hub genes in the progression of RILI-MMP9, IL-1ß, CCR1 and S100A9. The expression levels of the hub genes were verified in RILI mouse models, with S100A9 showing the highest level of overexpression. The level of S100A9 in bronchoalveolar lavage fluid (BALF) and the expression of S100A9 in lung tissues were positively correlated with the degree of inflammation in RILI. The results above indicate that S100A9 is a potential biomarker for the early prediction and diagnosis of the development of RILI.

Exosome-Related FTCD Facilitates M1 Macrophage Polarization and Impacts the Prognosis of Hepatocellular Carcinoma.

BACKGROUND: Exosomes are essential for hepatocellular carcinoma (HCC) progression and have garnered significant interest as novel targets for diagnostic, prognostic, and therapeutic approaches. This study aims to identify potential exosome-related biomarkers for the development of useful strategies for HCC diagnosis and therapy. METHODS: Three datasets obtained from the Gene Expression Omnibus (GEO) were utilized to identify differentially expressed genes (DEGs) in HCC. Through Gene Ontology (GO) analysis and protein-protein interaction (PPI) network, overall survival (OS) analysis, Cox analyses, and diethylnitrosamine (DEN)-induced HCC mouse model detection, exosome-related hub gene was screened out, followed by a prognostic value assessment and immune-correlates analysis based on the Cancer Genome Atlas (TCGA) dataset. The hub gene-containing exosomes derived from Hepa1-6 cells were isolated and characterized using differential ultracentrifugation, transmission electron microscopy scanning, and Western blot. Ultrasound-guided intrahepatic injection, cell co-culture, CCK-8, and flow cytometry were performed to investigate the effects of the hub gene on macrophage infiltration and polarization in HCC. RESULTS: A total of 83 DEGs enriched in the extracellular exosome term, among which, FTCD, HRA, and C8B showed the strongest association with the progression of HCC. FTCD was independently associated with a protective effect in HCC and selected as the hub gene. The presence of FTCD in exosomes was confirmed. FTCD-stimulated macrophages were polarized towards the M1 type and suppressed HCC cells proliferation. CONCLUSIONS: FTCD is a potential exosome-related biomarker for HCC diagnosis, prognosis, and treatment. The crosstalk between FTCD-containing exosomes and macrophages in HCC progression deserves further investigation.

Separation of Antioxidants from Trace Fraction of Ribes himalense via Chromatographic Strategy and Their Antioxidant Activity Supported with Molecular Simulations.

Antioxidants from natural sources have long been of interest to researchers. In this paper, taking the traditional Tibetan medicine Ribes himalense as an example, an integrated approach was used to identify and isolate its chemical composition with free-radical-scavenging properties from its ethanol extract. First, the ethanol extract of Ribes himalense was pretreated using polyamide medium-pressure liquid chromatography (polyamide-MPLC), and the target fraction (Fr4) was obtained. Then, a combined HPLC mode was utilized to purify antioxidants in Fr4 under the guidance of an online HPLC-1,1-diphenyl-2-picrylhydrazyl (HPLC-DPPH) activity screening system. Finally, three antioxidants (3-caffeoylquinic acid methyl ester, rutin, and myricetin-3'-α-L-rhamnopyranoside) were isolated, and this is the first report of their presence in R. himalense. Further molecular docking studies showed that the antioxidants exhibited good binding with HO-1, Nrf2, and iNOS. In conclusion, this comprehensive approach is capable of extracting high-purity antioxidants from trace fractions of Ribes himalense and holds promise for future applications in the exploration of the chemical compositions and bioactivity of natural products.

Jogging and weight training associated with increased high-density lipoprotein cholesterol levels in Taiwanese adults.

Background: Although previous studies have shown that aerobic and resistance exercise increase high-density lipoprotein cholesterol (HDL-C) levels, the optimal type of exercise has not been determined. Therefore, the purpose of this study was to investigate the association of jogging (a type of aerobic exercise) and weight training (a type of resistance exercise) with HDL-C levels in Taiwanese adults. Methods: The data used in this cross-sectional study were obtained from the Taiwan Biobank (TWB), which is a national health resource that contains the genetic information of Taiwanese volunteers aged 30-70 years. A total of 75,635 subjects (47,881 women and 27,754 men) were included in this study. The subjects were divided into four groups: jogging (n = 2,278), weight training (n = 522), mixed exercise (n = 519), and no exercise (n = 72,316). The TWB data were collected through questionnaires (e.g. basic characteristics, lifestyle factors, and disease history), biochemical tests, and anthropometric measurements. Results: Compared with no exercise, jogging, weight training, and mixed exercise were all associated with higher HDL-C levels (ß = 2.5470, 2.6249, and 3.2117, respectively). As seen, the ß value was highest for the mixed exercise group, followed by weight training and then jogging (p for trend <0.0001). Conclusions: In the current study, jogging and weight training were individually associated with higher levels of HDL-C. Engaging in both activities was associated with much higher levels of HDL-C. Our findings suggest that regular jogging and weight training might play an important role in increasing HDL-C levels.

Deep Learning-Based Automatic Detection of ASPECTS in Acute Ischemic Stroke: Improving Stroke Assessment on CT Scans.

(1) Background: The Alberta Stroke Program Early CT Score (ASPECTS) is a standardized scoring tool used to evaluate the severity of acute ischemic stroke (AIS) on non-contrast CT (NCCT). Our aim in this study was to automate ASPECTS. (2) Methods: We utilized a total of 258 patient images with suspected AIS symptoms. Expert ASPECTS readings on NCCT were used as ground truths. A deep learning-based automatic detection (DLAD) algorithm was developed for automated ASPECTS scoring based on 168 training patient images using a convolutional neural network (CNN) architecture. An additional 90 testing patient images were used to evaluate the performance of the DLAD algorithm, which was then compared with ASPECTS readings on NCCT as performed by physicians. (3) Results: The sensitivity, specificity, and accuracy of DLAD for the prediction of ASPECTS were 65%, 82%, and 80%, respectively. These results demonstrate that the DLAD algorithm was not inferior to radiologist-read ASPECTS on NCCT. With the assistance of DLAD, the individual sensitivity of the ER physician, neurologist, and radiologist improved. (4) Conclusion: The proposed DLAD algorithm exhibits a reasonable ability for ASPECTS scoring on NCCT images in patients presenting with AIS symptoms. The DLAD algorithm could be a valuable tool to improve and accelerate the decision-making process of front-line physicians.

Associations between cigarette smoking status and health-related physical fitness performance in male Taiwanese adults.

Background: The highest proportion of smoking behavior occurs in male adults in Taiwan. However, to our knowledge, no study has investigated the relationship between smoking behavior and health-related physical fitness according to education level, health status, betel nut-chewing status and obesity in male adults aged 18 years or older in Taiwan. Aims: This study aimed to determine the associations between cigarette smoking and health-related physical fitness performance in male Taiwanese adults. Methods: This was a cross-sectional study conducted on 27,908 male adults (aged 23-64 years) who participated in Taiwan's National Physical Fitness Survey 2014-2015. Data from a standardized structured questionnaire, anthropometric variables, and health-related physical fitness measurements were analyzed. Individuals were categorized as never smoking cigarettes, former smoker, and current smoker. Multiple linear regression analysis was performed to evaluate the association between cigarette smoking and health-related physical fitness performance. Results: Never smoking group exhibited a lower (p < 0.05) proportion of abdominal obesity, higher (p < 0.05) proportion of perceived good health status, and greater (p < 0.05) performance in 1-min sit-up and sit-and-reach tests when compared with current smoking and former smoking group. Former smoking group had the highest (p < 0.05) performance in 3-min step test among all groups. Current smoker was significantly negatively (p < 0.05) associated with 3-min step, 1-min sit-up and sit-and-reach tests. Notably, former smoker was significantly positively (p < 0.05) associated with 3-min step and 1-min sit-up tests, but still negatively (p < 0.05) associated with sit-and-reach performance. Conclusion: Current smoker was associated with an increased the risk of abdominal obesity, reduced the perceived health status and health-related physical fitness performance. Quitting smoking had beneficial effect on the perceived good health status, cardiorespiratory and muscular fitness in male Taiwanese adults, but not on flexibility performance. Further research on the ameliorate mechanism underlying this phenomenon is warranted.

Enantioselective Biosynthesis of L-Phenyllactic Acid From Phenylpyruvic Acid In Vitro by L-Lactate Dehydrogenase Coupling With Glucose Dehydrogenase.

As a valuable versatile building block, L-phenyllactic acid (L-PLA) has numerous applications in the fields of agriculture, pharmaceuticals, and biodegradable plastics. However, both normally chemically synthesized and naturally occurring PLA are racemic, and the production titer of L-PLA is not satisfactory. To improve L-PLA production and reduce the high cost of NADH, an in vitro coenzyme regeneration system of NADH was achieved using the glucose dehydrogenase variant LsGDHD255C and introduced into the L-PLA production process. Here an NADH-dependent L-lactate dehydrogenase-encoding variant gene (L-Lcldh1Q88A/I229A) was expressed in Pichia pastoris GS115. The specific activity of L-LcLDH1Q88A/I229A (Pp) was as high as 447.6 U/mg at the optimum temperature and pH of 40°C and 5.0, which was 38.26-fold higher than that of wild-type L-LcLDH1 (Pp). The catalytic efficiency (k cat/K m) of L-LcLDH1Q88A/I229A (Pp) was 94.3 mM-1 s-1, which was 67.4- and 25.5-fold higher than that of L-LcLDH1(Pp) and L-LcLDH1Q88A/I229A (Ec) expressed in Escherichia coli, respectively. Optimum reactions of L-PLA production by dual-enzyme catalysis were at 40°C and pH 5.0 with 10.0 U/ml L-LcLDH1Q88A/I229A (Pp) and 4.0 U/ml LsGDHD255C. Using 0.1 mM NAD+, 400 mM (65.66 g/L) phenylpyruvic acid was completely hydrolyzed by fed-batch process within 6 h, affording L-PLA with 90.0% yield and over 99.9% ee p. This work would be a promising technical strategy for the preparation of L-PLA at an industrial scale.

Gram-Scale Synthesis of (R)-P-Chlorophenyl-1,2-Ethanediol at High Concentration by a Pair of Epoxide Hydrolases.

(R)-p-chlorophenyl-1,2-ethanediol (pCPED) is an important intermediate for the synthesis of (R)-eliprodil that is widely applied in the treatment of ischemic stroke. To prepare (R)-pCPED with high enantiomeric excess (ee p) and yield via the enantioconvergent hydrolysis of racemic styrene oxide (rac-pCSO) at high concentration, the bi-enzymatic catalysis was designed and investigated by a pair of epoxide hydrolases, a mutant (PvEH1Z4X4-59) of Phaseolus vulgaris EH1 and a mutant (RpEHF361V) of Rhodotorula paludigena RpEH. Firstly, the maximum allowable concentration of rac-pCSO was confirmed. Subsequently, the addition mode and the weight ratio of two Escherichia coli cells were optimized. Finally, under the optimized reaction conditions-the cell weight ratio 20:1 of E. coli/pveh1z4x4-59 to E. coli/rpeh F361V, a simultaneous addition mode, and reaction temperature at 25°C-300 mM rac-pCSO in the 100 ml 4% (v/v) Tween-20/phosphate buffer system (100 mM, pH 7.0) was completely hydrolyzed within 5 h, affording (R)-pCPED with 87.8% ee p, 93.4% yield, and 8.63 g/L/h space-time yield (STY). This work would be an efficient technical strategy for the preparation of chiral vicinal diols at industrial scale.

Structure-Guided Regulation in the Enantioselectivity of an Epoxide Hydrolase to Produce Enantiomeric Monosubstituted Epoxides and Vicinal Diols via Kinetic Resolution.

Structure-guided microtuning of an Aspergillus usamii epoxide hydrolase was executed. One mutant, A214C/A250I, displayed a 12.6-fold enhanced enantiomeric ratio (E = 202) toward rac-styrene oxide, achieving its nearly perfect kinetic resolution at 0.8 M in pure water or 1.6 M in n-hexanol/water. Several other beneficial mutants also displayed significantly improved E values, offering promising biocatalysts to access 19 structurally diverse chiral monosubstituted epoxides (97.1 - ≥ 99% ees) and vicinal diols (56.2-98.0% eep) with high yields.

Structure-guided improvement in the enantioselectivity of an Aspergillus usamii epoxide hydrolase for the gram-scale kinetic resolution of ortho-trifluoromethyl styrene oxide.

Microtuning the substrate-binding pocket (SBP) of EHs has emerged as an effective approach to manipulate their enantio- or regio-selectivities and activities towards target substrates. Here, the enantioselectivity (enantiomeric ratio, E) of AuEH2 towards a racemic (rac-) ortho-trifluoromethyl styrene oxide (o-TFMSO) was improved via microtuning its SBP. Based on the analysis on the crystal structure of AuEH2, its specific residues I192, Y216, R322 and L344 lining the SBP in close to the catalytic triad were identified for site-saturation mutagenesis. After screening, five single-site mutants were selected with E values elevated from 8 to 12-25 towards rac-o-TFMSO. To further improve E, four double-site mutants were constructed by combinatorial mutagenesis of AuEH2R322V separately with AuEH2I192V, AuEH2Y216F, AuEH2L344A and AuEH2L344C. Among all the mutants, AuEH2R322V/L344C possessed the largest E of 83 with activity of 67 U/g wet cell. The kinetic resolution of 200 mM rac-o-TFMSO was conducted at 0 °C for 5.5 h using 80 mg/mL wet cells of E. coli/Aueh2R322V/L344C, a transformant expressing AuEH2R322V/L344C, retaining (S)-o-TFMSO with 98.4 % ees and 49.3 % yields. Furthermore, the molecular docking simulation analysis indicated that AuEH2R322V/L344C more enantiopreferentially attacks the terminal carbon (Cß) in the oxirane ring of (R)-o-TFMSO than AuEH2.

The effect of redox potential on the removal characteristic of divalent cations during activated carbon-based capacitive deionization.

The main objective of the study is to explore the removal characteristics of Cu2+ and Zn2+ ions in activated carbon-based capacitive deionization (CDI). In this work, CDI experiments were performed to remove divalent ions (e.g., Cu2+, Zn2+, and Ca2+) from single- and multicomponent aqueous solutions. As evidenced, divalent heavy metals could be successfully removed by charging the CDI cell at 1.2 V. Notably, the preferential removal of Cu2+ ions over Zn2+ and Ca2+ ions was observed in the charging step. The removal capacities for Cu2+, Zn2+, and Ca2+ ions in a competitive environment were 29.6, 19.6, and 13.8 µmol/g, respectively. In contrast, the regeneration efficiencies for the removal of Cu2+ and Zn2+ were much lower than that of Ca2+, suggesting the occurrence of irreversible Faradaic reactions on the cathode. X-ray photoelectron spectroscopy analysis demonstrated that Cu2+ ions were reduced to Cu(I) and Zn2+ ions were transformed to ZnO/Zn(OH)2 on the cathode. Therefore, there were two major mechanisms for the removal of divalent heavy metal ions: capacitive electrosorption and cathodic electrodeposition. Specifically, the reduction potential played a crucial role in determining the removal characteristics. When regarding divalent cations with similar hydrated sizes, the divalent cation with a higher reduction potential tended to be separated by cathodic electrodeposition rather than double-layer charging, indicating the high removal selectivity of activated carbon-based CDI. This paper constitutes a significant contribution to promoting the application of CDI for contaminant sequestration.

Improvement in the catalytic performance of a phenylpyruvate reductase from Lactobacillus plantarum by site-directed and saturation mutagenesis based on the computer-aided design.

To enhance the specific activity and catalytic efficiency (k cat/K m) of an NADH-dependent LpPPR, its directed modification was performed based on the computer-aided design using molecular docking simulation and multiple sequence alignment. Firstly, five single-site variants of an LpPPR-encoding gene (lpppr) were amplified and expressed in E. coli BL21 (DE3). The asymmetric reduction of 20 mM phenylpyruvic acid (PPA) was carried out using 50 mg/mL E. coli/lpppr R53Q or /lpppr A79V whole wet cells at 37 °C for 20 min, giving d-phenyllactic acid (PLA) with 41.1 or 44.3% yield, being 1.17- or 1.26-fold that by E. coli/lpppr. Secondly, double-site variants were obtained by saturation mutagenesis of Ala79 in LpPPRR53Q. Among all tested E. coli transformants, E. coli/lpppr R53Q/A79V exhibited the highest d-PLA yield of 85.3%. The specific activity and k cat/K m of the purified LpPPRR53Q/A79V increased to 67.5 U/mg and 169.8 mM-1 s-1, which were 3.0- and 13.2-fold those of LpPPR, respectively. Finally, the catalytic mechanism analysis of LpPPRR53Q/A79V by molecular docking simulation indicated that the replacement of Arg53 in LpPPR with Gln expanded its substrate-binding pocket, while that Ala79 with Val formed an additional π-sigma interaction with phenyl group of PPA. SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s13205-020-02633-3) contains supplementary material, which is available to authorized users.