scholarly journals A Novel L-Asparaginase from Hyperthermophilic Archaeon Thermococcus sibiricus: Heterologous Expression and Characterization for Biotechnology Application

2021 ◽  
Vol 22 (18) ◽  
pp. 9894
Author(s):  
Maria Dumina ◽  
Alexander Zhgun ◽  
Marina Pokrovskaya ◽  
Svetlana Aleksandrova ◽  
Dmitry Zhdanov ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Food Industry ◽  
Elevated Temperatures ◽  
Specific Activity ◽  
Human Fibroblasts ◽  
Initial Activity ◽  
Promising Candidate ◽  
Hyperthermophilic Archaeon ◽  
Normal Human ◽  
Biotechnology Application

L-asparaginase (L-ASNase) is a vital enzyme with a broad range of applications in medicine and food industry. Drawbacks of current commercial L-ASNases stimulate the search for better-producing sources of the enzyme, and extremophiles are especially attractive in this view. In this study, a novel L-asparaginase originating from the hyperthermophilic archaeon Thermococcus sibiricus (TsA) was expressed in Escherichia coli, purified and characterized. The enzyme is optimally active at 90 °C and pH 9.0 with a specific activity of 2164 U/mg towards L-asparagine. Kinetic parameters KM and Vmax for the enzyme are 2.8 mM and 1200 µM/min, respectively. TsA is stable in urea solutions 0–6 M and displays no significant changes of the activity in the presence of metal ions Ni2+, Cu2+, Mg2+, Zn2+ and Ca2+ and EDTA added in concentrations 1 and 10 mmol/L except for Fe3+. The enzyme retains 86% of its initial activity after 20 min incubation at 90 °C, which should be enough to reduce acrylamide formation in foods processed at elevated temperatures. TsA displays strong cytotoxic activity toward cancer cell lines K562, A549 and Sk-Br-3, while normal human fibroblasts WI-38 are almost unsensitive to it. The enzyme seems to be a promising candidate for further investigation and biotechnology application.

An Enzyme Linked Immunosorbent Assay for Determination of Tissue Plasminogen Activator Applied to Patients with Thromboembolic Disease

1983 ◽  
Vol 50 (03) ◽  
pp. 740-744 ◽  
Author(s):  
Nils Bergsdorf ◽  
Torbjörn Nilsson ◽  
Per Wallén
Keyword(s):  
Plasminogen Activator ◽  
Tissue Plasminogen Activator ◽  
Specific Activity ◽  
Thromboembolic Disease ◽  
Tissue Type ◽  
Enzyme Linked Immunosorbent Assay ◽  
Type Plasminogen Activator ◽  
Tissue Plasminogen ◽  
Normal Human

SummaryUtilizing the immunoglobulin fraction from a goat antiserum against human uterine tissue plasminogen activator, an enzyme- linked immunoassay for tissue-type plasminogen activator in human plasma has been developed. With the new method, the concentration of t-PA in normal human acidified plasma is found to be 4.0 ± 1.8 (SD) ng/ml. It increases to 12 ng/ml after a tomiquet test, and to 14 ng/ml after strenous physical exercise. In a group of patients with idiopathic thromboembolic disease, the resting t-PA concentration was 5 ng/ml and the post-occlusion value 16 ng/ml. Furthermore, the patients also exhibited a normal post-occlusion rise in the concentration of plasmin-α2-antiplasmin complex. However, in 37% of the post-occlusion patient plasmas, virtually no increase in t-PA could be detected by a specific activity assay. The results indicate that the reason for a defective post-occlusion fibrinolytic activity in a majority of cases may be the presence of increased concentrations of a fast-acting specific t-PA inhibitor.

Specialized systems for the processing of mnemonic information within the primate frontal cortex

1996 ◽  
Vol 351 (1346) ◽  
pp. 1455-1462 ◽  
Keyword(s):  
Frontal Cortex ◽  
Specific Activity ◽  
Human Subjects ◽  
Long Term Memory ◽  
Executive Processing ◽  
Severe Impairment ◽  
Normal Human ◽  
Short And Long Term ◽  
Cortical Association Areas

The lateral frontal cortex is involved in various aspects of executive processing within short- and long-term memory. It is argued that the different parts of the lateral frontal cortex make distinct contributions to memory that differ in terms of the level of executive processing that is carried out in interaction with posterior cortical systems. According to this hypothesis, the mid-dorsolateral frontal cortex (areas 46 and 9) is a specialized system for the monitoring and manipulation of information within working memory, whereas the mid-ventrolateral frontal cortex (areas 47/12 and 45) is involved in the active retrieval of information from the posterior cortical association areas. Data are presented which support this two-level hypothesis that posits two distinct levels of interaction of the lateral frontal cortex with posterior cortical association areas. Functional activation studies with normal human subjects have demonstrated specific activity within the mid-dorsolateral region of the frontal cortex during the performance of tasks requiring monitoring of self-generated and externally generated sequences of responses. In the monkey, lesions restricted to this region of the frontal cortex yield a severe impairment in performance of the above tasks, this impairment appearing against a background of normal performance on several basic mnemonic tasks. By contrast, a more severe impairment follows damage to the mid-ventrolateral frontal region and functional activation studies have demonstrated specific changes in activity in this region in relation to the active retrieval of information from memory.

scholarly journals Precursor-Boosted Production of Metabolites in Nasturtium officinale Microshoots Grown in Plantform Bioreactors, and Antioxidant and Antimicrobial Activities of Biomass Extracts

Molecules ◽  
2021 ◽  
Vol 26 (15) ◽  
pp. 4660
Author(s):  
Marta Klimek-Szczykutowicz ◽  
Michał Dziurka ◽  
Ivica Blažević ◽  
Azra Đulović ◽  
Małgorzata Miazga-Karska ◽  
...  
Keyword(s):  
Photosynthetic Pigments ◽  
Human Fibroblasts ◽  
Antimicrobial Activities ◽  
Precursor Feeding ◽  
Nasturtium Officinale ◽  
Qualitative And Quantitative ◽  
Normal Human ◽  
Inhibition Zones ◽  
Ferulic Acids ◽  
Antioxidant And Antimicrobial Activities

The study demonstrated the effects of precursor feeding on the production of glucosinolates (GSLs), flavonoids, polyphenols, saccharides, and photosynthetic pigments in Nasturtium officinale microshoot cultures grown in Plantform bioreactors. It also evaluated the antioxidant and antimicrobial activities of extracts. L-phenylalanine (Phe) and L-tryptophan (Trp) as precursors were tested at 0.05, 0.1, 0.5, 1.0, and 3.0 mM. They were added at the beginning (day 0) or on day 10 of the culture. Microshoots were harvested after 20 days. Microshoots treated with 3.0 mM Phe (day 0) had the highest total GSL content (269.20 mg/100 g DW). The qualitative and quantitative profiles of the GSLs (UHPLC-DAD-MS/MS) were influenced by precursor feeding. Phe at 3.0 mM stimulated the best production of 4-methoxyglucobrassicin (149.99 mg/100 g DW) and gluconasturtiin (36.17 mg/100 g DW). Total flavonoids increased to a maximum of 1364.38 mg/100 g DW with 3.0 mM Phe (day 0), and polyphenols to a maximum of 1062.76 mg/100 g DW with 3.0 mM Trp (day 0). The precursors also increased the amounts of p-coumaric and ferulic acids, and rutoside, and generally increased the production of active photosynthetic pigments. Antioxidant potential increased the most with 0.1 mM Phe (day 0) (CUPRAC, FRAP), and with 0.5 mM Trp (day 10) (DPPH). The extracts of microshoots treated with 3.0 mM Phe (day 0) showed the most promising bacteriostatic activity against microaerobic Gram-positive acne strains (MIC 250–500 µg/mL, 20–21 mm inhibition zones). No extract was cytotoxic to normal human fibroblasts over the tested concentration range (up to 250 μg/mL).

scholarly journals Generation of nonviral integration-free human iPS cell line KISCOi001-A from normal human fibroblasts, under defined xeno-free and feeder-free conditions

2021 ◽  
Vol 51 ◽  
pp. 102193
Author(s):  
Jose Inzunza ◽  
Jonathan Arias-Fuenzalida ◽  
Juan Segura-Aguilar ◽  
Ivan Nalvarte ◽  
Mukesh Varshney
Keyword(s):  
Cell Line ◽  
Human Fibroblasts ◽  
Ips Cell ◽  
Normal Human

scholarly journals Gut Digestive Function and Microbiome after Correction of Experimental Dysbiosis in Rats by Indigenous Bifidobacteria

2021 ◽  
Vol 9 (3) ◽  
pp. 522
Author(s):  
Lyudmila V. Gromova ◽  
Elena I. Ermolenko ◽  
Anastasiya L. Sepp ◽  
Yulia V. Dmitrieva ◽  
Anna S. Alekseeva ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Digestive Enzymes ◽  
Specific Activity ◽  
Control Group ◽  
Enteropathogenic Escherichia Coli ◽  
Beneficial Bacteria ◽  
Opportunistic Bacteria ◽  
Dyspeptic Symptoms ◽  
Specific Activities ◽  
Impaired Motor Function

In recent years, great interest has arisen in the use of autoprobiotics (indigenous bacteria isolated from the organism and introduced into the same organism after growing). This study aimed to evaluate the effects of indigenous bifidobacteria on intestinal microbiota and digestive enzymes in a rat model of antibiotic-associated dysbiosis. Our results showed that indigenous bifidobacteria (the Bf group) accelerate the disappearance of dyspeptic symptoms in rats and prevent an increase in chyme mass in the upper intestine compared to the group without autoprobiotics (the C1 group), but significantly increase the mass of chyme in the colon compared to the C1 group and the control group (healthy animals). In the Bf group in the gut microbiota, the content of opportunistic bacteria (Proteus spp., enteropathogenic Escherichia coli) decreased, and the content of some beneficial bacteria (Bifidobacterium spp., Dorea spp., Blautia spp., the genus Ruminococcus, Prevotella, Oscillospira) changed compared to the control group. Unlike the C1 group, in the Bf group there was no decrease in the specific activities of maltase and alkaline phosphatase in the mucosa of the upper intestine, but the specific activity of maltase was decreased in the colon chyme compared to the control and C1 groups. In the Bf group, the specific activity of aminopeptidase N was reduced in the duodenum mucosa and the colon chyme compared to the control group. We concluded that indigenous bifidobacteria can protect the microbiota and intestinal digestive enzymes in the intestine from disorders caused by dysbiosis; however, there may be impaired motor function of the colon.

scholarly journals Scaffold Hopping of α-Rubromycin Enables Direct Access to FDA-Approved Cromoglicic Acid as a SARS-CoV-2 MPro Inhibitor

2021 ◽  
Vol 14 (6) ◽  
pp. 541
Author(s):  
Hani A. Alhadrami ◽  
Ahmed M. Sayed ◽  
Heba Al-Khatabi ◽  
Nabil A. Alhakamy ◽  
Mostafa E. Rateb
Keyword(s):  
Binding Free Energy ◽  
Human Fibroblasts ◽  
Critical Time ◽  
Dynamics Simulation ◽  
Direct Access ◽  
Energy Calculation ◽  
Wide Range ◽  
Normal Human ◽  
Novel Scaffold

The COVID-19 pandemic is still active around the globe despite the newly introduced vaccines. Hence, finding effective medications or repurposing available ones could offer great help during this serious situation. During our anti-COVID-19 investigation of microbial natural products (MNPs), we came across α-rubromycin, an antibiotic derived from Streptomyces collinus ATCC19743, which was able to suppress the catalytic activity (IC50 = 5.4 µM and Ki = 3.22 µM) of one of the viral key enzymes (i.e., MPro). However, it showed high cytotoxicity toward normal human fibroblasts (CC50 = 16.7 µM). To reduce the cytotoxicity of this microbial metabolite, we utilized a number of in silico tools (ensemble docking, molecular dynamics simulation, binding free energy calculation) to propose a novel scaffold having the main pharmacophoric features to inhibit MPro with better drug-like properties and reduced/minimal toxicity. Nevertheless, reaching this novel scaffold synthetically is a time-consuming process, particularly at this critical time. Instead, this scaffold was used as a template to explore similar molecules among the FDA-approved medications that share its main pharmacophoric features with the aid of pharmacophore-based virtual screening software. As a result, cromoglicic acid (aka cromolyn) was found to be the best hit, which, upon in vitro MPro testing, was 4.5 times more potent (IC50 = 1.1 µM and Ki = 0.68 µM) than α-rubromycin, with minimal cytotoxicity toward normal human fibroblasts (CC50 > 100 µM). This report highlights the potential of MNPs in providing unprecedented scaffolds with a wide range of therapeutic efficacy. It also revealed the importance of cheminformatics tools in speeding up the drug discovery process, which is extremely important in such a critical situation.

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