• Title/Summary/Keyword: Microbial enzymes

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The Analysis and Application of a Recombinant Monooxygenase Library as a Biocatalyst for the Baeyer- Villiger Reaction

  • Park, Ji-Yeoun;Kim, Dong-Hyun;Kim, Su-Jin;Kim, Jin-Hee;Bae, Ki-Hwan;Lee, Choong-Hwan
    • Journal of Microbiology and Biotechnology
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    • v.17 no.7
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    • pp.1083-1089
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    • 2007
  • Because of their selectivity and catalytic efficiency, BVMOs are highly valuable biocatalysts for the chemoenzymatic synthesis of a broad range of useful compounds. In this study, we investigated the microbial Baeyer-Villiger oxidation and sulfoxidation of thioanisole and bicyclo[3.2.0]hept-2-en-6-one using whole Escherichia coli cells that recombined with each of the Baeyer-Villiger monooxygenases originated from Pseudomonas aeruginosa PAOl and two from Streptomyces coelicolor A3(2). The three BVMOs were identified in the microbial genome database by a recently described protein sequence motif; e.g., BVMO motif(FXGXXXHXXXW). The reaction products were identified as (R)-/(S)-sulfoxide and 2-oxabicyclo/3-oxabicyclo[3.3.0]oct-6-en-2-one by GC-MS analysis. Consequently, this study demonstrated that the three enzymes can indeed catalyze the Baeyer-Villiger reaction as a biocatalyst, and effective annotation tools can be efficiently exploited as a source of novel BVMOs.

Effective Production of N-Acetyl-$\beta$-glucosamine by Serratia marcescens Using Chitinadceous Waste

  • Kim, Kwang;A. Louise Creagh;Charles A. Haynes
    • Biotechnology and Bioprocess Engineering:BBE
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    • v.3 no.2
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    • pp.71-77
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    • 1998
  • The strain of Serratia marcescens QM B1466 produces selectively large amount of chitinolytic enzymes (about 1mg/L medium). Enzymatic hydrolysis of chitin to N-acetyl-${\beta}$-D-glucosamine (NAG) was performed with a system consisting of two hydrolases (chitinase and chitobiase) produced by optimization of a microbial host consuming chitin particles. For the development of Large-scale biological process for the production of NAG from chitinaceous waste, the selection and optimization of a microbial host, particle size of crab/shrimp chitin sources and initial induction time using chitin as a sole carbon source on chitinase/chitobiase production and NAG production were examined. Crab-shell chitin(1.5%) treated by dilute acid and , ball-milled with a normal diameter less than 250m gave the highest chitinase activity over a 7 days culture. Crude chitinase/ chitobiase solution obtained in a 10 L fed-batch fermentation showed a maximum activities of 23.6 U/mL and 5.1 U/mL, respectively with a feeding time of 3 hrs, near pH 8.5 at 30$^{\circ}C$.

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Protein Engineering of Deoxynucleoside Kinase from Lactobacillus acidophilus: Effect of Site-Directed Mutagenesis on Microbial Growth

  • Park, Inshik;Kim, Eun-Ae;Bang, Keuk-Seung;Kim, Seok-Hwan;Kim, Gi-Nahm;Lee, Min-Kyung;Kil, Ji-Oeun
    • Preventive Nutrition and Food Science
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    • v.6 no.1
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    • pp.79-81
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    • 2001
  • Deoxynucleoside kinases exist as heterodimeric pairs specific for deoxyadenosine/deoxyguanosine kinase (dAK/dGK) and deoxyadenosine/deoxycytidine kinase (dAK/dCK). The aspartic acid-84 in dGK was mutated to alanine, asparagine and glutamic acid by site-directed mutagenesis. The mutation resulted in a drastic decease in dGK activity compared to the unmodified cloned enzyme while it increased production of dAK activity. The mutated dak/dgk genes, which synthesize tandem deoxyadenosine/deoxyguanosine kinase, were inserted back to the Lactobacillus acidophilus and Lactococcus lactis by electroporation to determine the effect of site-directed mutation of he enzymes on the microbial growth. However, no significant change was observed in cell growth and lactic acid production between wild type and mutant lactic acid bacteria.

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Development of Postharvest Technologies to Preserve High Quality of Greenhouse Horticultural Commodities and their Processed Products -Development of Natural Antimicrobial Agents for preserving Greenhouse Fresh Produce(II) (남부지역 시설원예산물의 저장성 향상 및 가공품 개발에 관한 연구-시설원예산물의 선도유지를 위한 항균소재의 개발을 중심으로(II)-)

  • 조성환;정순경;김영록
    • Food Science and Preservation
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    • v.7 no.1
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    • pp.1-7
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    • 2000
  • To develop natural antimicrobial agents for keeping qualities of postharvested greenhouse produce the antimiocrobial actions of Polygonum cuspidatum Sieb. et Zucc. extract , which showed remarkable antimicrobial effects against microorganism causing the postharvest decay of greenhouse produce, were investigate. In the inhibitory experiment of enzymes related to energy production metabolism hexokinase activities decreased to 73% and 68% by treating with Polygonum cuspidatum Sieb. et.Zucc. extract and Eugenia caryophyllata Thumnberg extract in comparison with control, respectively. Direct visualization of microbial cells by using both transmission electron microscope and scanning electron microscope showed that microbial cell membrane was destroyed by treating with the dilute extract solution. this change of celluloar membrane permeability could be identified in the experiment that 0-nitrophenyl-${\beta}$-D-galactopyrano-side(ONPG), the artificial substrate of ${\beta}$-galactosidase, was hydrolyzed in the presence of the extract, indicating that the membrane was perturbed. The separation and identification of the most antimicrobialo substances isolated from Polygonum cuspidatum Sieb et. Zucc. extract and Eugenia caryophyllata Thunberg extract were carried out by using gas chromatography and mass spectrometry 9GC/MSD), which were identified as eugenol. As a result, the functionality of Polygonum cuspidatum Sieb. et Zucc. extract and Eugenia caryophyllata Thunberg extract as antimicrobial agents for keeping qualities of postharvested greenhouse produce may be recommended.

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Pharmacological Viewpoint Concerns and Phytochemical Components of Ginseng

  • RANA, Anvi
    • The Korean Journal of Food & Health Convergence
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    • v.8 no.5
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    • pp.21-28
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    • 2022
  • Ginseng is described as the "King of all herbs, "Man-root" or "Root of heaven" and regarded as the most powerful herbal remedy, particularly grown in Korea, China, Japan, Vietnam, and North America. It has been in existence for a long time. The most demanded herbal cure, Ginseng, principally the root, has long been employed in traditional Asian medicine. The extent of availability of bioactive combinations and their impact on the body differs between American and Asian ginseng. Asian ginseng, also known as Panax ginseng, has a more calming influence and is more advantageous than American ginseng, such as Panax quinquefolius. The pharmaceutical aspect of development and extraction with diverse morphological properties is examined. Saponins, glycosides, carbohydrates, polyacetylenes, amino acids, vitamins, volatile oil, enzymes are all present in the Phyto-content of Ginseng. Ginsenosides are saponins that are constituents of the triterpenoid dammarane and have anticancer, anti-cardiovascular, anti-microbial, anti-obesity, anti-inflammatory, and antioxidant properties. Ginseng, in particular, has the possibility to help with microbial invasion, inflammatory processes, oxidative stress, and diabetes. It developed nanoparticles and nanocomposite film technologies as novel drug delivery platforms for cancer, inflammation, and neurological illnesses. Furthermore, it offers a range of applications that will be vital for future growth.

Improved Biosurfactant Production by Bacillus subtilis SPB1 Mutant Obtained by Random Mutagenesis and Its Application in Enhanced Oil Recovery in a Sand System

  • Bouassida, Mouna;Ghazala, Imen;Ellouze-Chaabouni, Semia;Ghribi, Dhouha
    • Journal of Microbiology and Biotechnology
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    • v.28 no.1
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    • pp.95-104
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    • 2018
  • Biosurfactants or microbial surfactants are surface-active biomolecules that are produced by a variety of microorganisms. Biodegradability and low toxicity have led to the intensification of scientific studies on a wide range of industrial applications for biosurfactants in the field of environmental bioremediation as well as the petroleum industry and enhanced oil recovery. However, the major issues in biosurfactant production are high production cost and low yield. Improving the bioindustrial production processes relies on many strategies, such as the use of cheap raw materials, the optimization of medium-culture conditions, and selecting hyperproducing strains. The present work aims to obtain a mutant with higher biosurfactant production through applying mutagenesis on Bacillus subtilis SPB1 using a combination of UV irradiation and nitrous acid treatment. Following mutagenesis and screening on blood agar and subsequent formation of halos, the mutated strains were examined for emulsifying activity of their culture broth. A mutant designated B. subtilis M2 was selected as it produced biosurfactant at twice higher concentration than the parent strain. The potential of this biosurfactant for industrial uses was shown by studying its stability to environmental stresses such as pH and temperature and its applicability in the oil recovery process. It was practically stable at high temperature and at a wide range of pH, and it recovered above 90% of motor oil adsorbed to a sand sample.

Characterization of Two Metagenome-Derived Esterases That Reactivate Chloramphenicol by Counteracting Chloramphenicol Acetyltransferase

  • Tao, Weixin;Lee, Myung-Hwan;Yoon, Mi-Young;Kim, Jin-Cheol;Malhotra, Shweta;Wu, Jing;Hwang, Eul-Chul;Lee, Seon-Woo
    • Journal of Microbiology and Biotechnology
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    • v.21 no.12
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    • pp.1203-1210
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    • 2011
  • Function-driven metagenomic analysis is a powerful approach to screening for novel biocatalysts. In this study, we investigated lipolytic enzymes selected from an alluvial soil metagenomic library, and identified two novel esterases, EstDL26 and EstDL136. EstDL26 and EstDL136 reactivated chloramphenicol from its acetyl derivates by counteracting the chloramphenicol acetyltransferase (CAT) activity in Escherichia coli. These two enzymes showed only 27% identity in amino acid sequence to each other; however both preferentially hydrolyzed short-chain p-nitrophenyl esters (${\leq}C_5$) and showed mesophilic properties. In vitro, EstDL136 catalyzed the deacetylation of 1- and 3-acetyl and 1,3-diacetyl derivates; in contrast, EstDL26 was not capable of the deacetylation at $C_1$, indicating a potential regioselectivity. EstDL26 and EstDL136 were similar to microbial hormone-sensitive lipase (HSL), and since chloramphenicol acetate esterase (CAE) activity was detected from two other soil esterases in the HSL family, this suggests a distribution of CAE among the soil microorganisms. The isolation and characterization of EstDL26 and EstDL136 in this study may be helpful in understanding the diversity of CAE enzymes and their potential role in releasing active chloramphenicol in the producing bacteria.

Sugarcane Bagasse Hydrolysis Using Yeast Cellulolytic Enzymes

  • de Souza, Angelica Cristina;Carvalho, Fernanda Paula;Silva e Batista, Cristina Ferreira;Schwan, Rosane Freitas;Dias, Disney Ribeiro
    • Journal of Microbiology and Biotechnology
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    • v.23 no.10
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    • pp.1403-1412
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    • 2013
  • Ethanol fuel production from lignocellulosic biomass is emerging as one of the most important technologies for sustainable development. To use this biomass, it is necessary to circumvent the physical and chemical barriers presented by the cohesive combination of the main biomass components, which hinders the hydrolysis of cellulose and hemicellulose into fermentable sugars. This study evaluated the hydrolytic capacity of enzymes produced by yeasts, isolated from the soils of the Brazilian Cerrado biome (savannah) and the Amazon region, on sugarcane bagasse pre-treated with $H_2SO_4$. Among the 103 and 214 yeast isolates from the Minas Gerais Cerrado and the Amazon regions, 18 (17.47%) and 11 (5.14%) isolates, respectively, were cellulase-producing. Cryptococcus laurentii was prevalent and produced significant ${\beta}$-glucosidase levels, which were higher than the endo- and exoglucanase activities. In natura sugarcane bagasse was pre-treated with 2% $H_2SO_4$ for 30 min at $150^{\circ}C$. Subsequently, the obtained fibrous residue was subjected to hydrolysis using the Cryptococcus laurentii yeast enzyme extract for 72 h. This enzyme extract promoted the conversion of approximately 32% of the cellulose, of which 2.4% was glucose, after the enzymatic hydrolysis reaction, suggesting that C. laurentii is a good ${\beta}$-glucosidase producer. The results presented in this study highlight the importance of isolating microbial strains that produce enzymes of biotechnological interest, given their extensive application in biofuel production.

Effects of Tween 80 and Fibrolytic Enzymes on Ruminal Fermentation and Digestibility of Feeds in Holstein Cows

  • Baah, J.;Shelford, J.A.;Hristov, A.N.;McAllister, T.A.;Cheng, K.J.
    • Asian-Australasian Journal of Animal Sciences
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    • v.18 no.6
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    • pp.816-824
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    • 2005
  • The effects of the nonionic surfactant Tween 80 and a mixture of fibrolytic enzymes on total tract digestion, in situ disappearance (ISD) and ruminal fermentation characteristics of orchardgrass hay and barley grain were investigated in a 4${\times}$4 Latin square experiment with 4 non-lactating Holstein cows and 4 diets in 4 periods. Cows were offered a total mixed ration of 50% rolled barley grain and 50% orchardgrass hay treated with either 1) water (control), 2) 0.2% (vol/wt) Tween 80, 3) 0.2% (vol/wt) hydrolytic enzyme, or 4) 0.2% hydrolytic enzyme plus 0.2% Tween 80. Total tract digestibility coefficients of DM, nitrogen, NDF and ADF were not affected (p>0.05) by dietary treatment. Compared to the control, the rate of ISD of DM from orchardgrass hay was faster (p<0.05) in cows receiving diets treated with the enzyme alone or with enzyme plus Tween 80 (0.06/h vs. 0.076 and 0.069/h). The rate of digestion was lower (p<0.05) as compared to control when barley grain was treated with these additives. Ruminal fluid pH and concentrations of total VFA, acetate, isobutyrate and butyrate were not affected (p>0.05) by treatments. Cows that consumed diets treated with enzyme plus Tween 80 had higher (p<0.05) ruminal concentrations of propionate and isovalerate, and lower (p<0.05) acetate:propionate ratios. Compared to the control, microbial protein synthesis tended (p = 0.13) to increase with the addition of enzyme to the diet while nonammonia nitrogen flow to the duodenum increased (p<0.05) with both enzyme and Tween 80 treatments. The study indicated that fibrolytic enzymes alone or in combination with Tween 80 could enhance ISD of orchardgrass hay and ruminal concentrations of propionate, valerate and iso-valerate, but do not affect total tract digestibility.

Effect of Diet on Enzyme Profile, Biochemical Changes and In sacco Degradability of Feeds in the Rumen of Buffalo

  • Kamra, D.N.;Saha, Sudipto;Bhatt, Neeru;Chaudhary, L. C.;Agarwal, Neeta
    • Asian-Australasian Journal of Animal Sciences
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    • v.16 no.3
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    • pp.374-379
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    • 2003
  • Four rumen fistulated Murrah buffaloes were used to study the effect of four diets differing in roughage to concentrate ratio on rumen biochemical changes, microbial enzyme profile and in sacco degradability of feed in a $4{\times}4$ Latin Square design. The animals were fed four diets consisting of 80:20, 70:30, 60:40 and 50:50 ratios of wheat straw and concentrate mixtures, respectively. Wheat straw and concentrate mixture were mixed with water (0.6 l/kg feed) and complete feed mixture was offered to the animals at 8:00 h and 16:00 h in two equal parts. The variation in pH of rumen liquor (difference of maximum and minimum during 0-8 h post feeding) increased with increasing level of concentrate mixture in the diet. There was no effect of diet composition on volatile fatty acids, total nitrogen and trichloro-acetic acid precipitable nitrogen in the rumen liquor, but ammonia nitrogen increased with increasing level of concentrate mixture in the ration. Major portions of all fibre degrading enzymes were present in the particulate material (PM) of the rumen contents, but protease was absent in PM fraction. The activities of micro-crystalline cellulase, acetyl esterase and protease increased with increase in the level of concentrate mixture, but the activities of other enzymes (carboxymethylcellulase, filter paper degrading activity, xylanase, $\beta$-glucosidase and $\beta$-xylosidase) were not affected. The in sacco degradability and effective degradability of feeds increased with increasing level of concentrate mixture in the ration.