Dyerbowden3931

Laccases or benzenediol oxygen oxidoreductases (EC 1.10.3.2) are polyphenol multicopper oxidases that are known for their structural and functional diversity in various life forms. In the present study, the molecular and physico-chemical properties (redox-potential and secondary structures) of fungal laccase isozymes (FLIs) isolated from a medicinal mushroom Ganoderma lucidum were analyzed and compared with those of the recombinant bacterial laccases (rLac) obtained from different Yersinia enterocolitica strains. It was revealed that the FLIs contained His-Cys-His as the most conserved residue in its domain I Cu site, while the fourth and fifth residues were variable (Ile, Leu, or Phe). Evidently, the cyclic voltammetric measurements of Glac L2 at Type 1 Cu site revealed greater E° for ABTS/ABTS+ (0.312 V) and ABTS+/ABTS2+ (0.773 V) compared to the E° of rLac. Furthermore, circular dichroism-based conformational analysis revealed structural stability of the FLIs at acidic pH (3.0) and low temperature (70 °C). The zymographic studies further confirmed the functional inactivation of FLIs at high temperatures (≥70 °C), predominantly due to domain unfolding. These findings provide novel insight into the evolution of the catalytic efficiency and redox properties of the FLIs, contributing to the existing knowledge regarding stress responses, metabolite production, and the biotechnological utilization of metabolites.Accumulating evidence indicates that plant cell wall-associated receptor-like kinases (WAKs) involve in defense against pathogen attack, but their related signaling processes and regulatory mechanism remain largely unknown. We identified a WAK-like kinase (GhWAKL) from upland cotton (Gossypium hirsutum) and characterized its functional mechanism. Expression of GhWAKL in cotton plants was induced by Verticillium dahliae infection and responded to the application of salicylic acid (SA). Knockdown of GhWAKL expression results in the reduction of SA content and suppresses the SA-mediated defense response, enhancing cotton plants susceptibility to V. dahliae. And, ecotopic overexpression of GhWAKL in Arabidopsis thaliana conferred plant resistance to the pathogen. Further analysis demonstrated that GhWAKL interacted with a cotton DnaJ protein (GhDNAJ1) on the cell membrane. Silencing GhDNAJ1 also enhanced cotton susceptibility to V. dahliae. NG25 datasheet Moreover, the mutation of GhWAKL at site Ser628 with the phosphorylation decreased the interaction with GhDNAJ1 and compromised the plant resistance to V. dahliae. We propose that GhWAKL is a potential molecular target for improving resistance to Verticillium wilt in cotton.Mesorhizobium loti carbonic anhydrase (MlCA), an intrinsically high catalytic enzyme, has been employed for carbon dioxide capture and sequestration. However, recombinant expression of MlCA in Escherichia coli often forms inclusion bodies. Hence, protein partners such as fusion-tags and molecular chaperones are involved in regarding reduce the harshness of protein folding. TrxA-tag and GroELS have been chosen to co-express with MlCA in E. coli under an inducible T7 promoter or a constitutive J23100 promoter to compare productivity and activity. The results possessed that coupling protein partners effectively increased soluble MlCA up to 2.9-folds under T7 promoter, thus enhancing the CA activity by 120% and achieving a 5.2-folds turnover rate. Besides, it has also shifted the optimum temperature from 40 °C to 50 °C, promoted stability in the broad pH range (4.5 to 9.5) and the presence of various metal ions. Based on the in vitro assay and isothermal titration calorimetry (ITC) analysis, GroELS enhancing CA activity was due to change the intrinsic thermodynamic properties of the enzyme from endothermic to exothermic reaction (i.e., ∆H = 89.8 to -121.8 kJ/mol). Therefore, the collaboration of TrxA-MlCA with GroELS successfully augmented CO2 biomineralization.Solid-state is the preferred choice for storage of protein therapeutics to improve stability and preserve the biological activity by decreasing the physical and chemical degradation associated with liquid protein formulations. Lyophilization or freeze-drying is an effective drying method to overcome the instability problems of proteins. However, the processing steps (freezing, primary drying and secondary drying) involved in the lyophilization process can expose the proteins to various stress and harsh conditions, leading to denaturation, aggregation often a loss in activity of protein therapeutics. Stabilizers such as sugars and surfactants are often added to protect the proteins against physical stress associated with lyophilization process and storage conditions. Another way to curtail the degradation of proteins due to process related stress is by modification of the lyophilization process. Slow freezing, high nucleation temperature, decreasing the extent of supercooling, and annealing can minimize the formation of the interface (ice-water) by producing large ice crystals with less surface area, thereby preserving the native structure and stability of the proteins. Hence, a thorough understanding of formulation composition, lyophilization process parameters and the choice of analytical methods to characterize and monitor the protein instability is crucial for development of stable therapeutic protein products. This review provides an overview of various stress conditions that proteins might encounter during lyophilization process, mechanisms to improve the stability and analytical techniques to tackle the proteins instability during both freeze-drying and storage.Herein, the immobilization of α-amylase onto hydroxyapatite (HA) and hydroxyapatite-decorated ZrO2 (10%wt) (HA-ZrO2) nanocomposite were investigated. The immobilization yield was 69.7% and 84% respectively. The structural differences were characterized using X-Ray diffraction, attenuated total reflectance-Fourier transform infrared spectra, Raman, and scanning electron microscope. After 10 repeated cycles, the residual activity of immobilized α-amylase onto HA and HA-ZrO2 nanocomposite was 46% and 70%, respectively. The storage stability was recorded to be 27%, 50% and 69% from its initial activity in the case of free and immobilized enzyme onto HA and HA-ZrO2 nanocomposite, respectively after 8 weeks. The pH-activity profile and temperature revealed pH 6.0 and temperature 50 °C as the optimal values of free α-amylase, while the optimum values for α-amylase on HA and HA-ZrO2 was shifted to pH 6.5 and 60 °C after immobilization. The immobilized α-amylase onto HA-ZrO2 showed comparatively higher catalytic activity than the free α-amylase. The Km value after the immobilization process onto HA was 2.1 folds highly than that of the free enzyme. In conclusion, it can be inferred that HA-ZrO2 is more sustainable and beneficial support for enzyme immobilization and it represents promising supports for different uses of α-amylase in the biomedical applications.α-Amylase inhibitors (α-AIs) delay digestion of dietary starch by inhibiting α-amylase in the gut, thereby reducing the postprandial glycemia, which is beneficial to the patients with obesity and diabetes. The proteinaceous α-AIs from wheat can effectively control starch digestion and regulate postprandial hyperglycemia. However, their gastric intolerance remains a challenge, which limits its commercial production and industrial application. In this study, sodium alginate/chitosan aerogels loaded with wheat protein α-AIs were prepared and evaluated as potential transportation and protection matrices for important components in food or pharmaceutical applications. Specifically, the biodegradable aerogel cross-linked with sodium alginate-chitosan-calcium chloride, has a large surface area and open porous structure, which can adsorb staple wheat proteins as an integrated edible material to block around 88,660 U/g of α-amylase activity. The aerogel particles were able to protect the activity of wheat α-AIs in the stomach, leading to the slow passage of the wheat α-AIs through the small intestine to inhibit starch digestion more effectively. Animal experiments further showed that the postprandial blood glucose levels in rats were effectively controlled through delayed increase, after administration of wheat protein-functionalized aerogel particles loaded with wheat α-AIs, which are natural biological macromolecules. This is a novel, safe, and economical method for the prevention and pretreatment of diabetes.The Hsp90 chaperone system interacts with a wide spectrum of client proteins, forming variable and dynamic multiprotein complexes that involve the intervention of cochaperone partners. Recent results suggest that the role of Hsp90 complexes is to establish interactions that suppress unwanted client activities, allow clients to be protected from degradation and respond to biochemical signals. Cryo-electron microscopy (cryoEM) provided the first key molecular picture of Hsp90 in complex with a kinase, Cdk4, and a cochaperone, Cdc37. Here, we use a combination of molecular dynamics (MD) simulations and advanced comparative analysis methods to elucidate key aspects of the functional dynamics of the complex, with different nucleotides bound at the N-terminal Domain of Hsp90. The results reveal that nucleotide-dependent structural modulations reverberate in a striking asymmetry of the dynamics of Hsp90 and identify specific patterns of long-range coordination between the nucleotide binding site, the client binding pocket, the cochaperone and the client. Our model establishes a direct atomic-resolution cross-talk between the ATP-binding site, the client region that is to be remodeled and the surfaces of the Cdc37-cochaperone.While variants of noncoding RNAs (ncRNAs) have been experimentally validated as a new class of biomarkers and drug targets, the discovery and interpretation of relationships between ncRNA variants and human diseases become important and challenging. Here we present ncRNAVar (http//www.liwzlab.cn/ncrnavar/), the first database that provides association data between validated ncRNA variants and human diseases through manual curation on 2650 publications and computational annotation. ncRNAVar contains 4565 associations between 711 human disease phenotypes and 3112 variants from 2597 ncRNAs. Each association was reviewed by professional curators, incorporated with valuable annotation and cross references, and designated with an association score by our refined score model. ncRNAVar offers web applications including association prioritization, network visualization, and relationship mapping. ncRNAVar, presenting a landscape of ncRNA variants in human diseases and a useful resource for subsequent software development, will improve our insight of relationships between ncRNA variants and human health.Adolescence is associated with behavioral changes offering opportunities for prevention of cardiovascular risk behaviors. Primary care physicians are ideally placed to deliver preventive interventions to adolescents. The objective was to systematically review the evidence about effectiveness of primary care-led interventions addressing the main cardiovascular risk behaviors in adolescents physical activity, sedentary behaviors, diet and smoking. PubMed, Embase, PsycINFO, CINAHL, Cochrane, ClinicalTrials.gov, and ISRCTN registry were searched from January 1990 to April 2020. Randomized controlled trials of interventions in primary care contexts on at least one of the cardiovascular behaviors were included, targeting 10-19-year old adolescents, according to the World Health Organization's definition. Two authors independently assessed risk of bias. Twenty-two papers were included in the narrative synthesis, reporting on 18 different studies. Interventions targeting smoking uptake seemed more effective than interventions targeting established smoking or the three other risk behaviors.