Process conditions optimization and study of pH and thermal stability of free and immobilized gutaminase-free recombinant L-asparaginase II of Pactobacterium carotovorum MTCC 1428 from Escherichia coli

2021 ◽  
Vol 16 (10) ◽  
pp. 63-74
Author(s):  
Rachna Goswami ◽  
Vijay Kumar Mishra ◽  
Veeranki Venkata Dasu
Keyword(s):  
Escherichia Coli ◽  
Food Processing ◽  
Immobilized Enzymes ◽  
Lymphocytic Leukemia ◽  
Process Conditions ◽  
Chitosan Beads ◽  
Optimum Ph ◽  
Temperature Levels ◽  
Glutaminase Activity ◽  
Thermal Stability Of

L-asparaginase (EC 3.5.1.1) is used for the treatment of acute lymphocytic leukemia and food processing of starch rich foods for reducing the acrylamide formation. In our current efforts, we have immobilized the purified glutaminase-free recombinant Lasparaginase II of Pectobacterium carotovorum MTCC 1428 from Escherichia coli BL21 (DE3) on glutaraldehyde activated chitosan beads. The purified recombinant L-asparaginase II has no partial glutaminase activity which is a pre-requisite to reduce the possibility of side effects during the course of anticancer therapy. In order to study the best conditions for the performance of free enzymes and immobilized enzymes, response surface methodology was used to optimize the pH and temperature of the process conditions. It was found that the most favorable pH and temperature for the free enzyme were pH 7.83 and 47.64°C while for the immobilized enzyme, the optimum pH and temperature levels were found to be 7.88 and 48.07 °C. Furthermore, the thermal and pH studies of free and immobilized enzymes were studied under various combinations of pH and temperature and finally the thermodynamic parameters of free and immobilized glutaminase-free recombinant asparaginase were evaluated.

scholarly journals Enhanced Wastewater Treatment by Immobilized Enzymes

2021 ◽  
Author(s):  
Jakub Zdarta ◽  
Katarzyna Jankowska ◽  
Karolina Bachosz ◽  
Oliwia Degórska ◽  
Karolina Kaźmierczak ◽  
...  
Keyword(s):  
Wastewater Treatment ◽  
Process Optimization ◽  
Organic Pollutants ◽  
Large Scale ◽  
Removal Rate ◽  
Immobilized Enzymes ◽  
Future Research ◽  
Process Conditions ◽  
Removal Rates ◽  
Catalytic Cycles

Abstract Purpose of Review In the presented review, we have summarized recent achievements on the use of immobilized oxidoreductases for biodegradation of hazardous organic pollutants including mainly dyes, pharmaceuticals, phenols, and bisphenols. In order to facilitate process optimization and achievement of high removal rates, effect of various process conditions on biodegradation has been highlighted and discussed. Recent Findings Current reports clearly show that immobilized oxidoreductases are capable of efficient conversion of organic pollutants, usually reaching over 90% of removal rate. Further, immobilized enzymes showed great recyclability potential, allowing their reuse in numerous of catalytic cycles. Summary Collected data clearly indicates immobilized oxidoreductases as an efficient biocatalytic tools for removal of hazardous phenolic compounds, making them a promising option for future water purification. Data shows, however, that both immobilization and biodegradation conditions affect conversion efficiency; therefore, process optimization is required to achieve high removal rates. Nevertheless, we have demonstrated future trends and highlighted several issues that have to be solved in the near-future research, to facilitate large-scale application of the immobilized oxidoreductases in wastewater treatment.

scholarly journals Purification and Characterization of Asparaginase fromPhaseolus vulgarisSeeds

2015 ◽  
Vol 2015 ◽  
pp. 1-6 ◽  
Author(s):  
Saleh A. Mohamed ◽  
Mohamed F. Elshal ◽  
Taha A. Kumosani ◽  
Alia M. Aldahlawi
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Lymphoblastic Leukemia ◽  
Potential Candidate ◽  
Purification And Characterization ◽  
Apparent Homogeneity ◽  
Wide Range ◽  
Optimum Ph ◽  
Glutaminase Activity ◽  
Low Activity

L-asparaginase from bacteria has been used in treatment of acute lymphoblastic leukemia. The aim of this study was to purify and characterize L-asparaginase fromPhaseolus vulgarisseeds instead of microbial sources. L-asparaginase was purified to apparent homogeneity. The enzyme has molecular mass of 79 kDa. The purified asparaginase had very low activity toward a number of asparagine and glutamine analogues. L-asparaginase was free from glutaminase activity. Kinetic parameters, Km andVmax of purified enzyme, were found to be 6.72 mM and 0.16 μM, respectively. The enzyme had optimum pH at 8.0. The enzyme showed high stability at alkaline pH (pH 7.5–9.0) when incubated for up to 24 h. L-asparaginase had the same temperature optimum and thermal stability at 37°C. K+was able to greatly enhance the activity of asparaginase by 150% compared with other metals tested. In conclusion, L-asparaginase showed no glutaminase activity and good stability over a wide range of physiological conditions, and thus it could be used as a potential candidate for treatment of acute lymphoblastic leukemia.

scholarly journals Searching for Antagonistic Activity of Bacterial Isolates Derived from Food Processing Environments on Some Food-Borne Pathogenic Bacteria

2020 ◽  
Vol 49 (4) ◽  
pp. 415-423
Author(s):  
B. Baráti-Deák ◽  
Cs. Mohácsi-Farkas ◽  
Á. Belák
Keyword(s):  
Escherichia Coli ◽  
Food Processing ◽  
Pathogenic Bacteria ◽  
Inhibitory Effect ◽  
Antagonistic Activity ◽  
Bacterial Isolates ◽  
Bacterial Strains ◽  
Food Borne ◽  
Time Period ◽  
Growth Decline

Bacterial strains with inhibitory effect on Salmonella Hartford, Listeria monocytogenes, Yersinia enterocolitica, and Escherichia coli, respectively, were isolated. Out of the 64 bacteria originated from food processing environments, 20 could inhibit at least one of the tested pathogens, and it was proved that growth decline of the pathogenic bacteria was more remarkable by co-culturing than by using cell-free supernatants of the isolates. Seven different genera (Pseudomonas, Bacillus, Paenibacillus, Macrococcus, Staphylococcus, Serratia, and Rothia) reduced the pathogens’ growth during the time period of analysis, and the strongest inhibitory effect was observed after 24 h between 15 and 30 °C. Sensitivity of the tested human pathogenic bacteria against the inhibitory strains was distinct, as Y. enterocolitica could be inhibited by numerous isolates, while S. Hartford proved to be the most resistant. Our results reveal that the isolated bacteria or their excreted metabolites could hinder pathogen growth when used in sufficient quantities.

Characterization of Thermal Stability of the Escherichia coli Fapy–DNA Glycosylase

2001 ◽  
Vol 288 (1) ◽  
pp. 121-128 ◽  
Author(s):  
Serguei V. Kuznetsov ◽  
Olga M. Sidorkina ◽  
Jacques Laval ◽  
Anjum Ansari
Keyword(s):  
Escherichia Coli ◽  
Thermal Stability ◽  
Dna Glycosylase ◽  
Thermal Stability Of

scholarly journals Facile Bioinspired Preparation of Fluorinase@Fluoridated Hydroxyapatite Nanoflowers for the Biosynthesis of 5′-Fluorodeoxy Adenosine

2020 ◽  
Vol 12 (1) ◽  
pp. 431
Author(s):  
Ningning Li ◽  
Bingjing Hu ◽  
Anming Wang ◽  
Huimin Li ◽  
Youcheng Yin ◽  
...  
Keyword(s):  
Synthetic Activity ◽  
Initial Activity ◽  
Organofluorine Compounds ◽  
X Ray Diffraction ◽  
Green Method ◽  
X Ray ◽  
Fluoridated Hydroxyapatite ◽  
Optimum Ph ◽  
Ft Ir ◽  
Thermal Stability Of

To develop an environmentally friendly biocatalyst for the efficient synthesis of organofluorine compounds, we prepared the enzyme@fluoridated hydroxyapatite nanoflowers (FHAp-NFs) using fluorinase expressed in Escherichia coli Rosetta (DE3) as the biomineralization framework. The obtained fluorinase@FHAp-NFs were characterized by scanning electron microscope (SEM), X-ray diffraction (XRD), and FT-IR spectrum and used in the enzymatic synthesis of 5′-fluorodeoxy adenosin with S-adenosyl-L-methionine and fluoride as substrate. At an optimum pH of 7.5, fluorinase confined in the hybrid nanoflowers presents an approximately 2-fold higher synthetic activity than free fluorinase. Additionally, after heating at 30 °C for 8 h, the FHAp-NFs retained approximately 80.0% of the initial activity. However, free enzyme could remain only 48.2% of its initial activity. The results indicate that the fluoride and hybrid nanoflowers efficiently enhance the catalytic activity and thermal stability of fluorinase in the synthesis of 5′-fluorodeoxy adenosine, which gives a green method for producing the fluorinated organic compounds.

scholarly journals 3D Chitin Scaffolds from the Marine Demosponge Aplysina archeri as a Support for Laccase Immobilization and Its Use in the Removal of Pharmaceuticals

Biomolecules ◽  
2020 ◽  
Vol 10 (4) ◽  
pp. 646 ◽  
Author(s):  
Jakub Zdarta ◽  
Tomasz Machałowski ◽  
Oliwia Degórska ◽  
Karolina Bachosz ◽  
Andriy Fursov ◽  
...  
Keyword(s):  
Storage Stability ◽  
Packed Bed ◽  
Packed Bed Reactor ◽  
Optimum Process ◽  
Process Conditions ◽  
Total Removal ◽  
Laccase Immobilization ◽  
Continuous Use ◽  
First Time ◽  
Thermal Stability Of

For the first time, 3D chitin scaffolds from the marine demosponge Aplysina archeri were used for adsorption and immobilization of laccase from Trametes versicolor. The resulting chitin–enzyme biocatalytic systems were applied in the removal of tetracycline. Effective enzyme immobilization was confirmed by scanning electron microscopy. Immobilization yield and kinetic parameters were investigated in detail, in addition to the activity of the enzyme after immobilization. The designed systems were further used for the removal of tetracycline under various process conditions. Optimum process conditions, enabling total removal of tetracycline from solutions at concentrations up to 1 mg/L, were found to be pH 5, temperature between 25 and 35 °C, and 1 h process duration. Due to the protective effect of the chitinous scaffolds and stabilization of the enzyme by multipoint attachment, the storage stability and thermal stability of the immobilized biomolecules were significantly improved as compared to the free enzyme. The produced biocatalytic systems also exhibited good reusability, as after 10 repeated uses they removed over 90% of tetracycline from solution. Finally, the immobilized laccase was used in a packed bed reactor for continuous removal of tetracycline, and enabled the removal of over 80% of the antibiotic after 24 h of continuous use.

scholarly journals Purification and characterization of the class-II D-fructose 1,6-bisphosphate aldolase from Escherichia coli (Crookes' strain)

1978 ◽  
Vol 169 (3) ◽  
pp. 633-641 ◽  
Author(s):  
S A Baldwin ◽  
R N Perham ◽  
D Stribling
Keyword(s):  
Escherichia Coli ◽  
Common Ancestor ◽  
Class Ii ◽  
Purification And Characterization ◽  
Thiol Compounds ◽  
E Coli ◽  
Optimum Ph ◽  
Class Ii Aldolase ◽  
Fructose 1,6 Bisphosphate

A new form of the class-II D-fructose 1,6-bisphosphate aldolase (EC 4.1.2.13) of Escherichia coli (Crookes' strain) was isolated from an extract of glycerol-grown bacteria. It has a higher molecular weight (approx. 80000)than previous preparations of the enzyme and closely resembles the typical class-II aldolase from yeast in size and amino acid composition. On the other hand, its kinetic behaviour is not typical of a class-II aldolase. The enzyme has no requirement for thiol compounds either for stability or activity, added K+ ions have no effect, and the optimum pH for the cleavage activity is unusually high. The class-II enzymes from the prokaryote E. coli and the eukaryote yeast show no immunological identity. However, the similarity of their structures suggests that they have evolved from a common ancestor.

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