Characterization Of Cellulase Research Articles

Purification and Characterization of Cellulase from <i>Aspergillus niger</i> Causing Soft Rot of White Yam in Three Environments in Nigeria

Aspergillus niger is abundant in most tropical soils and invariably, on the surface of yam tubers while still attached to the plant and on the root hairs during harvesting or storage. Naturally, the peridermic surface of the tubers function to exclude pathogen but damage caused by accidental incision or cut surface during weeding, insect attack and harvesting provide avenue for the infection. Aspergillus niger isolated from diseased yam in three zones in Nigeria synthesized cellulase which caused soft rot of the yam within nine days of inoculation. Microscopic and molecular analyses revealed two isolates of A. niger, P1 and P2, from different environment produced cellulase enzymes in significantly different proportions. When protein extracts from the infection were subjected to molecular exclusion chromatography, three peaks of absorption (A, B and C) were produced with only the components of peak A showing cellulase activity. Further fractionation of the components of peak A produced two absorption peaks (Aa and Ab) with only component Aa showing Cellulase activity. A. niger isolates, P1 and P2 showed considerable differences in the intensity of cellulase production suggesting that multiple strains of A. niger in the soil of yam-growing environments synthesized cellulase as transcriptional products in different manner underscoring the effect of physico – chemical properties of the soil on infectivity and virulence of the organism during yam rot.

Isolation of Cellulolytic Bacteria from Soil, Identification by 16S rRNA Gene Sequencing and Characterization of Cellulase

Microbial Cellulases have an escalating demand in many industries and constitute a major group of the industrial enzymes. It has attracted the attention of many researchers because of its tremendous industrial applications including textile industry, pulp, and paper industry, laundry and detergent industry, food and animal feed industry. The present study pursues to unfold a novel cellulase that can overcome existing challenges in biorefineries as well as to reduce biofuel production cost. Therefore, soil from a dairy farm was screened for potent cellulase producers on carboxymethylcellulose agar. Out of 68 isolates, 31 expressed cellulase activity. The best isolate so far had an extracellular crude enzyme activity of 0.167 U/ml and specific activity of 0.333 U/mg. The cell morphology, cultural characteristics, and biochemical tests presumptively identified it to belong to the genus Bacillus. Molecular analysis using 16S rRNA gene of the isolate indicated it to be Bacillus subtilis. The optimum pH and temperature for the activity of the crude enzyme were determined to be 5 and 65°C respectively.
 Bangladesh J Microbiol, Volume 33, Number 1-2, June-Dec 2016, pp 17-22

토양으로부터 단리한 Bacillus subtilis NSC 유래 Cellulase의 특성 규명

토양으로부터 단리한 Bacillus subtilis NSC 유래 Cellulase의 특성 규명

Isolation, Optimization and Characterization of Cellulases and Hemicellulases from Bacillus Cereus LAZ 518 Isolated from Cow Dung Using Corn Cobs as Lignocellulosic Waste

Isolation, Optimization and Characterization of Cellulases and Hemicellulases from Bacillus Cereus LAZ 518 Isolated from Cow Dung Using Corn Cobs as Lignocellulosic Waste

Characterization of cellulase from E. coli BPPTCC-EGRK2

Cellulase enzymes are widely used in various industries such as detergent industry, bioethanol, animal feed, textile and paper. This research focused on characterization of cellulase produced from Eschericia coli BPPT-CC EgRK2, which is a recombinant that can produce protein enzymes endo- β-1,4-glucanase. Eschericia coli BPPT-CC EgRK2 was cultured in 1 litre liquid medium Luria Bertani. Because the bacteria is intracellular, sonication is needed for cell disruption to get the cellulase enzyme. The enzyme activity was then analyzed by CMC substrate at different concentration. The protein content analysis was carried out using Bradford method; the molecular weight analysis was done using SDS-PAGE; while the enzyme kinetics was plotted using Michaelis-Menten model. Our results showed the highest enzyme activity was 2.403 U/ml and the protein concentration was 5.352 mg/ml. The Michaelis-Menten constant (Km) and maximum velocity (Vmax) for CMC substrate hydrolysis were 0.07 μmol/ml and 2.49 μmol/ml/sec, respectively. The cellulase molecular weight was 58 kDa using SDS-PAGE with 7.5% stacking gel. The results indicated that Eschericia coli BPPT-CC EgRK2 is a promising renewable source for cellulase production for industrial application.

삼림 토양으로부터 단리된 Bacillus sp. JB1 유래 Cellulase 특성 규명

삼림 토양으로부터 단리된 Bacillus sp. JB1 유래 Cellulase 특성 규명

Characterization of cellulase from Aspergillus tubingensis NKBP-55 for generation of fermentable sugars from agricultural residues

The aim of this work was to characterize cellulase from Aspergillus tubingensis NKBP-55 for generation of fermentable sugars from agricultural residues. The strain produced high titres of cellulase (750 U/gds) on copra meal in solid state fermentation (SSF). The enzyme preparation also showed hemicellulolytic activities (U/gds) viz. endo-mannanase (1023), endo-xylanase (167), β-glucosidase (72) and α-galactosidase (54). Zymography revealed presence of six cellulases, six mannanases and one β-glucosidase. It effectively degraded sugarcane bagasse (SCB) and rice straw (RS) releasing xylose, glucose and cellobiose. One cellulase (Cat 1, Mr ∼65 kDa) was purified and characterized. It retained more than 50% activity at 70 °C after 150 mins and its activity was enhanced in the presence of Mn2+ ions (130%) and β-mercaptoethanol (140%). FTIR and 13C CP/MAS NMR analysis of the enzyme treated SCB and RS revealed degradation of cellulose and hemicellulose, while 1H and 13C liquid state NMR experiments confirmed release of glucose.

토양에서 단리한 Bacillus sp. TH1 유래 Cellulase의 특성 규명

토양에서 단리한 Bacillus sp. TH1 유래 Cellulase의 특성 규명

Celulasas presentes en el tracto digestivo del camarón Litopenaeus vannamei

Las celulasas han sido clasificadas como endoglucanasas responsables de degradar celulosa, lo que le proporciona gran importancia en los procesos de la cadena alimentaria. Las celulasas y enzimas relacionadas son utilizadas ampliamente en varios procesos industriales. La celulosa es utilizada como una fuente nutricional por varios organismos, inicialmente se pensaba que únicamente microorganismos podrían degradarla, pero en la actualidad se ha encontrado celulasas en varios grupos de animales. En este trabajo, presentamos la detección de celulasas en extractos obtenidos del camarón Litopenaeus vannamei. En los ensayos se utilizaron individuos adultos y de los diferentes subestadios de L. vannamei colectados en los estanques de cría de las camaroneras de la Península de Santa Elena y Muisne, en las provincias de Santa Elena y Esmeraldas, respectivamente. Los hepatopáncreas, los estadios larvarios y los intestinos fueron homogeneizados en un homogeneizador Braun-Melsungen. Para la identificación de la celulasa, se utilizó como sustrato carboximetil celulosa (CMC) de mediana viscosidad. Para medir su actividad se añadió ácido 3,5 dinitrosalicílico (DNS). La actividad de la celulasa se determinó en base en la cantidad de los grupos reductores liberados. Para intentar determinar el origen sea endógeno o microbiano de las celulasas identificadas, se sometieron las muestras de adultos a antibióticos.
 
 En el subestadio zoea 3, se identificó una celulasa con un rango de pH óptimo entre pH 6.5 y pH 9.5. En las muestras de hepatopáncreas del camarón adulto se determinaron dos pH óptimos: uno en el rango ácido (pH 4-6) y otro en el rango básico (pH 10-11) lo cual sugiere la presencia de dos celulasas diferentes. En el intestino se observó actividad únicamente en el pH ácido (pH 4-6). La actividad celulolítica detectada en el desarrollo ontogénetico del camarón fue baja e inestable, una condición reportada también en el análisis comparativo de enzimas digestivas en otras cuatro especies de crustáceos. La actividad relativa de las dos enzimas del hepatopáncreas es similar y cada una de estas tienen una actividad cuatro veces mayor a la encontrada en el intestino. Estos resultados indican la presencia de dos celulasas presentes en el hepatopáncreas y se sugiere que una de ellas (con pH óptimo ácido) podría ser transportada desde el hepatopáncreas al intestino, aunque los datos no son concluyentes. En el caso de la celulasa con actividad a pH básico (10.5) puede tratarse de una celulasa producida de manera endógena por el camarón. Los datos reportados en este estudio sobre las capacidades de producción enzimática del camarón puede ser la base para futuros proyectos de investigación cuyo objetivo sea optimizar los procedimientos de cultivo del camarón.

Purification and characterization of cellulase from a novel isolate of Trichoderma longibrachiatum

ABSTRACTIn the present study, the isolation, purification and characterization of a complex enzyme, cellulase, was carried out using a new local isolate of the fungus Trichoderma longibrachiatum (KM274866) from wood chips, with a view to explore its utility in the biofuel industry. The fungus was grown on a selected natural substrate, sugarcane bagasse, based on cost considerations for enzyme production. The enzyme was purified 14.82 fold with a yield of 25.8% and specific activity of 30 U/mg of protein. The molecular mass of the enzyme was found to be 67 ± 1 KDa. The optimum pH was 4.8, but the enzyme was stable at a pH range of 3–6. Optimum temperature was 45 °C, but the stability range of the enzyme was 30–55 °C. Metal ions such as Ca2+, Na+, Mg2+, Zn2+ and Fe2+ enhanced enzyme activity. Triton X100 in the medium resulted in a significant increase of enzyme activity compared to other group specific reagents. KM and Vmax for the enzyme were found to be 0.121 mg/ml and 0.421 μmol/min, respectively, against the substrate carboxy methyl cellulose. As the enzyme is from an inexpensive source, it will be most useful in the preparation of bioethanol for the biofuel industry.

Production and characterization of cellulase from E. coli EgRK2 recombinant based oil palm empty fruit bunch

Oil Palm Empty Fruit Bunch (OPEFB) is an abundant biomass resource in Indonesia, which contains 41.3 ~ 46.5% (w/w) of cellulose. This research examined the production of cellulase by the E. coli EgRK2 recombinant strain using an OPEFB substrate. The production of the enzyme was initially examined to identify optimum growth conditions, by observing the growth and activity of E. coli EgRK2 compared to its wild type. Our results showed that the optimum production time, pH and temperature of the recombinant growth and cellulase activity were achieved at 24 h, and at 7 and 40°C, respectively. Using these optimum conditions, the enzyme was produced, and experiments were carried out to examine the enzyme characteristics, produced from both strains, on hydrolysis of cellulose from OPEFB. Our results showed that the activity of the enzyme produced by the recombinant almost doubled compared to that of the wild type, although the optimum pH for both strains was pH 6. Higher activity was achieved by the recombinant compared to the wild type strain, and values were 1.905 and 1.366 U/mL, respectively. The optimum temperature for hydrolysis by cellulase occurred at 50°C for Bacillus sp. RK2, and 60°C for Bacillus sp. EgRK2. The Michaelis-Menten constant (Km) and maximum velocity (Vmax) for OPEFB degradation by E. coli EgRK2 were 0.26% and 1.750 μmol/mL/sec, which were significantly better values than those of the wild type. Control experiments for the degradation test using CMC also showed a better Vmax value for E. coli EgRK2 compared to the wild type, which is 2.543 and 1.605 μmol/mL/sec, respectively.

토양 유래 Cellulase 생산균주의 Screening 및 Bacillus subtilis NCE3 유래 Cellulase 특성 규명

토양 유래 Cellulase 생산균주의 Screening 및 Bacillus subtilis NCE3 유래 Cellulase 특성 규명

Isolation, Identification and Characterization of Cellulase from a Bacterium Obtained at a Saw-Mill Site in Ile-Ife, Nigeria

Isolation, Identification and Characterization of Cellulase from a Bacterium Obtained at a Saw-Mill Site in Ile-Ife, Nigeria

Partial Purification and Characterization of Cellulase (Ec.3.2.1.4) from Aspergillus niger Using Saw Dust as Carbon Source

Cellulase (EC.3.2.1.4) was isolated and partially purified from the submerged culture of Aspergillus niger through a simple four step purification step using homogenization to obtain crude filtrate, ammonium sulphate precipitation, carboxymethyl cellulose chromatography, and sephadex G-200 chromatography. A purification fold of 1.00 and 1.26 were gotten for crude filtrate and ammonium sulphate precipitation respectively, while 0.56 and 0.41 were gotten for carboxymethyl cellulose and sephadex G-200. The highest yield of 96.4% was recorded for (NH4)2SO4 precipitation. Kinetic studies gave Km and Vmax of 0.01 mg/ml and 1.02 μmole/ml/min for carboxymethyl cellulose and 0.02 mg/ml and 0.96 μmole/ml/min for sephadex G-200. Optimum temperature and pH of 40°C and 8.0 were obtained for the cellulase. Ethanol was found to inhibit the activity of the enzyme higher than other inhibitors tested. The implication of this is that given a more suitable carbon source and a longer culturing time, a very useful cellulase with commercial value could be produced.

Purification and Characterization of Cellulase from Termite Ametermes eveuncifer (Silverstri) Soldiers

Cellulase enzyme was purified and characterized from termite soldiers (Ametermes eveuncifer) using 70% ammonium sulphate precipitation, ion exchange chromatography and affinity chromatography. The enzyme isolated had a specific activity of 5.04 U/mg with a percentage yield of 11.7%. The enzyme showed maximum activity at 500C and pH 8. The enzyme was not inhibited by Ba2+ at a concentration of 1mM and Pb2+ at 10 mM concentration but was inhibited by other metal ions at 1 mM and 10 mM concentrations of their salts (NaCl, KCl, MnCl2, and NiCl2),

Construction of recombinant sestc Saccharomyces cerevisiae for consolidated bioprocessing, cellulase characterization, and ethanol production by in situ fermentation.

Bioethanol is an important oil substitute produced by the sugar fermentation process. To improve the efficiency of cellulase expression of Saccharomyces cerevisiae, a eukaryotic expression vector harboring a single-enzyme-system-three-cellulase gene (sestc) was integrated into the S. cerevisiae genome by the protoplast method. Using PCR screening, RT-PCR, and “transparent circle” detection, several recombinant S. cerevisiae strains, capable of efficiently expressing the heterogeneous cellulase, were selected. The total activity of cellulase, endo-β-D-glucanase, exo-β-D-glucanase, and xylanase of the recombinant S. cerevisiae transformant (designated number 14) was 1.1, 378, 1.44, and 164 U ml−1, respectively, which was 27.5-, 63-, 24-, and 19-fold higher than that of the wild-type strain. The concentration of ethanol produced by the engineered S. cerevisiae strain was 8.1 gl−1, with wheat bran as the carbon source, under submerged conditions; this was 57.86-fold higher than that produced by the wild-type strain (0.14 gl−1).

Utilization of sea water based media for the production and characterization of cellulase by Fusarium subglutinans MTCC 11891

Abstract The present study focuses on isolation, screening and identification of fungal strain capable of producing cellulase enzyme in sea water based media from paddy fields of Rourkela, Odisha, India. The filamentous fungi isolated were identified as Fusarium subglutinans MTCC 11891. Cellulase enzyme was characterized for its optimal pH and temperature and also studied for the effect of metal ions on enzyme activity. Comparative studies were carried out using both fresh and sea water based media in order to investigate the salt tolerance level of cellulase produced by the fungal strain. The fungal strain was found to be halotolerant with optimal pH of 5.0 with cellulase activity of 292.53 U/mL and 184 U/mL in fresh and sea water based Mandel's media, respectively. The optimal temperature for F. subglutinans MTCC 11891 cellulase was recorded at 80 °C with 347.43 U/mL for fresh water and 232 U/mL for sea water based Mandel's media. Manifold increase in cellulase activity was evidenced in the presence of 5 mM Mn 2+ and Fe 2+ concentration in both fresh and sea water based Mandel's media suggesting these two cations as key catalytic molecules. Partial purification of the cellulase produced in fresh water based Mandel's media was performed using diethylaminoethanol (DEAE)-Sepharose column and the fraction with enzyme activity of 298.68 U/mL was recorded as fraction with highest cellulase concentration. Halotolerant cellulases would be more useful in future for the development of sea water based systems to produce bioethanol.

토양으로 단리된 Bacillus subtilis KB1 유래 Cellulase의 특성 규명 및 효소 유전자의 규명

토양으로 단리된 Bacillus subtilis KB1 유래 Cellulase의 특성 규명 및 효소 유전자의 규명

Screening, purification and characterization of cellulase from fungi isolated from used mushroom substrate

Alarge number of microorganism especially filamentous fungi has ability to degrade cellulose. The purpose of this study was to conduct screening, purification, and characterization of cellulase from fungi which was isolated from used paddy straw mushroom substrate. Screening of cellulytic activity using CMC medium shown that 11 out of 20 isolates of fungi produced a clearing zone surrounding fungal colony. Among them isolate number JMF 12 showed the highest cellulase activity and was further used for purification and characterization. The cellulase was purified to electrophoretical homogeneity by ammonium sulfate precipitation, dialyzed by Novagen D-Maxi TubeTM Dialyzer, and Sephadex G-100 gel filtration chromatography. The recovery and purification fold was 3.82 % and 1.98 respectively, after Sephadex G-100 gel filtration chromatograph. The o purified cellulase had an optimal pH and temperature at 6 and 45 C. The K m and V max of cellulase was 11.43 mM and 0.006mmol/min respectively. The purified cellulase was activated by Na + and Zn ++ but inhibited by Ca ++ , Co ++ , Fe ++ , and Hg ++ .

Enzyme-based lignocellulose hydrolyzation - Brief data survey for cellulase performance characterization on behalf of the Sauter mean diameter of raw material particles.

The data presented here supports the informational background of enzyme-based lignocellulose hydrolyzation, cellulase characterization, and sugar yield prediction for the work “Enzyme-based lignocellulose hydrolyzation – Sauter mean diameter of raw materials as a basis for cellulase performance characterization and yield prediction” by Glaser [1]. Glucose yields from the enzymatic hydrolysis of the raw materials were shown as a function of cellulase enzyme loading as well as of particle size with different solid loading. The data for the proposed methods of the determination of enzyme activity in inhomogeneous samples of lignocellulosic raw materials are presented. The data of the empirical model that was developed for the prediction of hydrolysis yields for different enzyme concentrations, substrate specific particle size, and solid loadings, are given. Data are also given in relation of terms of scale-up opportunities.