scholarly journals Biochemical Characterization of a Novel Bacterial Laccase and Improvement of Its Efficiency by Directed Evolution on Dye Degradation

2021 ◽  
Vol 12 ◽  
Author(s):  
Shuang Dai ◽  
Qian Yao ◽  
Gen Yu ◽  
Shan Liu ◽  
Jeonyun Yun ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Dye Degradation ◽  
Environmental Pollutants ◽  
Mutant Enzyme ◽  
Bromophenol Blue ◽  
Coomassie Brilliant Blue ◽  
Recombinant Laccase ◽  
Coomassie Brilliant ◽  
Acid Violet ◽  
Acid Violet 7

Laccase is a copper-containing polyphenol oxidase with a wide range of substrates, possessing a good application prospect in wastewater treatment and dye degradation. The purpose of this research is to study the degradation of various industrial dyes by recombinant laccase rlac1338 and the mutant enzyme lac2-9 with the highest enzyme activity after modification by error-prone PCR. Four enzyme activities improved mutant enzymes were obtained through preliminary screening and rescreening, of which lac2-9 has the highest enzyme activity. There are four mutation sites, including V281A, V281A, P309L, S318G, and D232V. The results showed that the expression of the optimized mutant enzyme also increased by 22 ± 2% compared to the unoptimized enzyme and the optimal reaction temperature of the mutant enzyme lac2-9 was 5°C higher than that of the rlac1338, and the optimal pH increased by 0.5 units. The thermal stability and pH stability of mutant enzyme lac2-9 were also improved. With ABTS as the substrate, the kcat/Km of rlac1338 and mutant strain lac2-9 are the largest than other substrates, 0.1638 and 0.618 s–1M–1, respectively, indicating that ABTS is the most suitable substrate for the recombinant enzyme and mutant enzyme. In addition, the Km of the mutant strain lac2-9 (76 μM) was significantly lower, but the kcat/Km (0.618 s–1M–1) was significantly higher, and the specific enzyme activity (79.8 U/mg) increased by 3.5 times compared with the recombinant laccase (22.8 U/mg). The dye degradation results showed that the use of rlac1338 and lac2-9 alone had no degradation effect on the industrial dyes [indigo, amaranth, bromophenol blue, acid violet 7, Congo red, coomassie brilliant blue (G250)], however, adding small molecular mediators Ca2+ and ABTS at the same time can significantly improve the degradation ability. Compared to the rlac1338, the degradation rates with the simultaneous addition of Ca2+ and ABTS of mutant enzyme lac2-9 for acid violet 7, bromophenol blue and coomassie brilliant blue significantly improved by 8.3; 3.4 and 3.4 times. Therefore, the results indicated that the error-prone PCR was a feasible method to improve the degradation activity of laccase for environmental pollutants, which provided a basis for the application of laccase on dye degradation and other environmental pollutants.

scholarly journals Biodegradation of Synthetic Dye by Endophytic Fungal Isolate in Calotropis procera Root

2015 ◽  
Vol 3 (3) ◽  
pp. 373-380 ◽  
Author(s):  
Shiv Kumar Verma ◽  
Anand Kumar ◽  
Moti Lal ◽  
Mira Debnath
Keyword(s):  
Methylene Blue ◽  
Rose Bengal ◽  
Dye Degradation ◽  
Fungal Endophytes ◽  
Brilliant Blue ◽  
Calotropis Procera ◽  
Methyl Red ◽  
Coomassie Brilliant Blue ◽  
Fungal Isolates ◽  
Coomassie Brilliant

In this study, based on colony morphology characteristics, a total of 19 fungal endophytes were isolated from root of Calotropis Procera a traditional Indian medicinal plant. All fungal isolates were screened for their dye degradation ability. The dyes used as test dyes were Rose Bengal (RB), azo dye Methyl Red (MR), Coomassie Brilliant Blue (CBB) and Methylene Blue (MB) and the concentration of each dye in the experiment was kept 100mg/L. Among the 19 fungal endophytic isolates (CPR1-CPR19), only one isolate CPR4 showed strong dye decolourization capability against all the four test dye. Dye decolourization ability by the isolate CPR4 was determined to be 97.4%, 87%, 65% and 45% for Rose Bengal (RB), Methyl Red (MR), Coomassie Brilliant Blue (CBB) and Methylene Blue (MB) respectively. Complete colour decolourization was observed with rose Bengal followed by Methyl Red. Glucose minimal medium was used for liquid and solid culture of fungal isolates. Fungal biomass production in the presence of four test dye was studied and compare with control culture of fungal endophytes. Effect of temperature, pH, stationary and agitation conditions on dye degradation was also studied.Int J Appl Sci Biotechnol, Vol 3(3): 373-380 

scholarly journals Partial purification and biochemical characterization of a new highly acidic NYSO laccase from Alcaligenes faecalis

2020 ◽  
Vol 18 (1) ◽  
Author(s):  
Soad A. Abdelgalil ◽  
Ahmad R. Attia ◽  
Reyed M. Reyed ◽  
Nadia A. Soliman
Keyword(s):  
Enzyme Activity ◽  
Sodium Dodecyl ◽  
Alcaligenes Faecalis ◽  
Maximum Activity ◽  
Sodium Oxalate ◽  
Partial Purification ◽  
Bromophenol Blue ◽  
Bromocresol Purple ◽  
Laccase Enzyme

Abstract Background Due to the multitude industrial applications of ligninolytic enzymes, their demands are increasing. Partial purification and intensive characterization of contemporary highly acidic laccase enzyme produced by an Egyptian local isolate designated Alcaligenes faecalis NYSO were studied in the present investigation. Results Alcaligenes faecalis NYSO laccase has been partially purified and intensively biochemically characterized. It was noticed that 40–60% ammonium sulfate saturation showed maximum activity. A protein band with an apparent molecular mass of ~ 50 kDa related to NYSO laccase was identified through SDS-PAGE and zymography. The partially purified enzyme exhibited maximum activity at 55 °C and pH suboptimal (2.5–5.0). Remarkable activation for enzyme activity was recognized after 10-min exposure to temperatures (T) 50, 60, and 70 °C; time elongation caused inactivation, where ~ 50% of activity was lost after a 7-h exposure to 60 °C. Some metal ions Cu2+, Zn2+, Co2+, Ni2+, Mn2+, Cd2+, Cr2+, and Mg2+ caused strong stimulation for enzyme activity, but Fe2+ and Hg2+ reduced the activity. One millimolar of chelating agents [ethylenediamine tetraacetic acid (EDTA), sodium citrate, and sodium oxalate] caused strong activation for enzyme activity. Sodium dodecyl sulfate (SDS), cysteine-HCl, dithiothreitol (DTT), β-mercaptoethanol, thioglycolic acid, and sodium azide caused strong inhibition for NYSO laccase activity even at low concentration. One millimolar of urea, imidazole, kojic acid, phenylmethylsulfonyl fluoride (PMSF), H2O2, and Triton X-100 caused activation. The partially purified NYSO laccase had decolorization activity towards different dyes such as congo red, crystal violet, methylene blue, fast green, basic fuchsin, bromophenol blue, malachite green, bromocresol purple eriochrome black T, and Coomassie Brilliant Blue R-250 with various degree of degradation. Also, it had a vast range of substrate specificity including lignin, but with high affinity towards p-anisidine. Conclusion The promising properties of the newly studied laccase enzyme from Alcaligenes faecalis NYSO strain would support several industries such as textile, food, and paper and open the possibility for commercial use in water treatment. It will also open the door to new applications due to its ligninolytic properties in the near future.

Total protein determined in human breast milk by use of coomassie brilliant blue and centrifugal analysis.

1989 ◽  
Vol 35 (10) ◽  
pp. 2127-2129 ◽  
Author(s):  
Y Bergqvist ◽  
L Karlsson ◽  
L Fohlin
Keyword(s):  
Breast Milk ◽  
Total Protein ◽  
Brilliant Blue ◽  
Human Breast Milk ◽  
Human Breast ◽  
Simple Method ◽  
Coomassie Brilliant Blue ◽  
Kjeldahl Method ◽  
Coomassie Brilliant

Abstract This simple method of centrifugal analysis for total protein in human breast milk is based on the change in the wavelength of the absorbance maximum of Coomassie Brilliant Blue G-250 when the dye is bound to protein. Within-run and between-day CVs were 3.8% and 4.8%, respectively. Compared with a micro-Kjeldahl method for determination of total nitrogen, the coefficient of correlation was 0.99.

Photocatalysis of Coomassie Brilliant Blue Using Clay Mineral

2016 ◽  
Vol 869 ◽  
pp. 765-767 ◽  
Author(s):  
Layane Rodrigues Almeida ◽  
João Sammy Nery Souza ◽  
Edson Cavalcanti Silva Filho ◽  
Josy Anteveli Osajima
Keyword(s):  
Organic Pollutants ◽  
Uv Light ◽  
Brilliant Blue ◽  
Blue Dye ◽  
Secondary Treatment ◽  
Toxic Compounds ◽  
Coomassie Brilliant Blue ◽  
Kinetics Of ◽  
Coomassie Brilliant ◽  
Photocatalytic Processes

The presence of organic pollutants, which cannot be eliminated by conventional processes of primary and secondary treatment, can be problematic. Photocatalytic processes offer an efficient breakdown of organic pollutants into non-toxic compounds such as CO2 and H2O. This paper proposes the use of the titanium dioxide embedded in palygorskite as a photoactive material in the degradation of cationic dye, Coomassie Brilliant Blue. The system was irradiated using UV light for a maximum time of 120 minutes. The concentration of the dye used was 1.0x10-4 mol L-1 in 0.5 g L-1 of the photoactive material. The kinetics of the system was monitored by UV-Vis spectrophotometry. In 120 minutes of radiation, the process of photocatalysis reduced the initial concentration of the Coomassie Brilliant Blue dye in half.

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