scholarly journals Purification and Characterization of Strong Simultaneous Enzyme Production of Protease and α-Amylase from an Extremophile-Bacillus sp. FW2 and Its Possibility in Food Waste Degradation

Fermentation ◽  
2021 ◽  
Vol 8 (1) ◽  
pp. 12
Author(s):  
Van Hong Thi Pham ◽  
Jaisoo Kim ◽  
Jeahong Shim ◽  
Soonwoong Chang ◽  
Woojin Chung
Keyword(s):  
Food Waste ◽  
Environmental Conditions ◽  
Enzyme Production ◽  
Molecular Weights ◽  
Sds Page ◽  
Metabolic Products ◽  
Wide Ph Range ◽  
Waste Degradation ◽  
Ph Range

Microbial enzymes such as protease and amylase are valuable enzymes with various applications, widely investigated for their applications in degradation of organic waste, biofuel industries, agricultural, pharmaceuticals, chemistry, and biotechnology. In particular, extremophiles play an important role in biorefinery due to their novel metabolic products such as high value catalytic enzymes that are active even under harsh environmental conditions. Due to their potentials and very broad activities, this study isolated, investigated, and characterized the protease- and amylase-producing bacterial strain FW2 that was isolated from food waste. Strain FW2 belongs to the genus Bacillus and was found to be closest to Bacillus amyloliquefaciens DSM 7T with a similarity of 99.86%. This strain was able to degrade organic compounds at temperatures from −6 °C to 75 °C (but weak at 80 °C) under a wide pH range (4.5–12) and high-salinity conditions up to 35% NaCl. Maximum enzyme production was obtained at 1200 ± 23.4 U/mL for protease and 2400 ± 45.8 U/mL for amylase for 4 days at pH 7–7.5, 40–45 °C, and 0–10% NaCl. SDS-PAGE analysis showed that the molecular weights of purified protease were 28 kDa and 44 kDa, corresponding to alkaline protease (AprM) and neutral protease (NprM), respectively, and molecular weight of α-amylase was 55 kDa. Degradation food waste was determined after 15 days, observing a 69% of volume decrease. A potential commercial extremozyme-producing bacteria such as strain FW2 may be a promising contributor to waste degradation under extreme environmental conditions.

scholarly journals Purification and Characterization of 2,6-β-d-Fructan 6-Levanbiohydrolase fromStreptomyces exfoliatus F3-2

2000 ◽  
Vol 66 (1) ◽  
pp. 252-256 ◽  
Author(s):  
Katsuichi Saito ◽  
Kazuya Kondo ◽  
Ichiro Kojima ◽  
Atsushi Yokota ◽  
Fusao Tomita
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Extracellular Enzyme ◽  
Molecular Weights ◽  
Sds Page ◽  
Monomeric Structure ◽  
Ph Range ◽  
Hydrolysis Of

ABSTRACT Streptomyces exfoliatus F3-2 produced an extracellular enzyme that converted levan, a β-2,6-linked fructan, into levanbiose. The enzyme was purified 50-fold from culture supernatant to give a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weights of this enzyme were 54,000 by SDS-PAGE and 60,000 by gel filtration, suggesting the monomeric structure of the enzyme. The isoelectric point of the enzyme was determined to be 4.7. The optimal pH and temperature of the enzyme for levan degradation were pH 5.5 and 60°C, respectively. The enzyme was stable in the pH range 3.5 to 8.0 and also up to 50°C. The enzyme gave levanbiose as a major degradation product from levan in an exo-acting manner. It was also found that this enzyme catalyzed hydrolysis of such fructooligosaccharides as 1-kestose, nystose, and 1-fructosylnystose by liberating fructose. Thus, this enzyme appeared to hydrolyze not only β-2,6-linkage of levan, but also β-2,1-linkage of fructooligosaccharides. From these data, the enzyme from S. exfoliatus F3-2 was identified as a novel 2,6-β-d-fructan 6-levanbiohydrolase (EC 3.2.1.64 ).

scholarly journals Production, Characterization of Tannase fromPenicillium montanenseURM 6286 under SSF Using Agroindustrial Wastes, and Application in the Clarification of Grape Juice (Vitis viniferaL.)

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Juliana Silva de Lima ◽  
Roberta Cruz ◽  
Julyanna Cordoville Fonseca ◽  
Erika Valente de Medeiros ◽  
Marília de Holanda Cavalcanti Maciel ◽  
...  
Keyword(s):  
Tannic Acid ◽  
Enzyme Production ◽  
Grape Juice ◽  
Tannin Content ◽  
Crude Enzyme Extract ◽  
Agroindustrial Waste ◽  
Wide Ph Range ◽  
Ph Range ◽  
Agroindustrial Wastes

Tannase is an enzyme that hydrolyzes esters and lateral bonds of tannins, such as tannic acid, releasing glucose and gallic acid and stands out in the clarification of wines and juices. Fungi of the generaAspergillusandPenicilliumare excellent producers of this enzyme. The search for fungi that produce high levels of tannase as well as new substrates for the enzyme production by the SSF is required. The objectives of this study were to evaluate the production of tannase byAspergillusandPenicilliumspecies through SSF using leaves and agroindustrial waste barbados cherry and mangaba fruit as substrate, select the best producer, optimize production, characterize the crude enzyme extract, and apply it the clarification of grape juice. Selecting the best producer was performed by planning Placket-Burman and RSM.P. montanenseshowed highest activity with 41.64 U/mL after 72 h of fermentation residue using barbados cherry, with 3.5% tannic acid and 70% moisture. The enzyme showed the highest activity at pH 9.0 and 50°C. The tannase ofP. montanensewas stable over a wide pH range and temperature and, when applied to grape juice, showed higher efficiency by reducing 46% of the tannin content after incubation 120 m.

Isolation and biochemical characterization of septate junctions: differences between the proteins in smooth and pleated varieties

1989 ◽  
Vol 93 (1) ◽  
pp. 123-131
Author(s):  
NANCY J. LANE ◽  
STEPHEN M. DILWORTH
Keyword(s):  
Periplaneta Americana ◽  
Biochemical Characterization ◽  
Septate Junction ◽  
Septate Junctions ◽  
Molecular Weights ◽  
Thin Sectioning ◽  
Sds Page ◽  
High Concentrations ◽  
Membrane Components

Septate junctions are found only in invertebrate tissues, and are almost ubiquitous within them. In arthropods, the two major types are the ‘pleated’ and the ‘smooth’ varieties. Using tissues from different species, including the cockroach Periplaneta americana, procedures have been established for obtaining membrane fractions selectively enriched in septate junctions. The junctions have been identified in pellets of these fractions by both thin sectioning and freeze-fracturing. SDS-PAGE of these membrane fractions reveals two major polypeptide species with apparent molecular weights of 22000–24000 and 17000–18000. Consistent differences in these apparent molecular weights are observed between the pleated and smooth varieties of septate junction. These polypeptides are probably integral membrane components, as they remain associated after treatment with high concentrations of urea. Evidence suggests a plane of weakness in the mid-line of the extracellular septal ribbons.

scholarly journals Characterization of a Robust and pH-Stable Tannase from Mangrove-Derived Yeast Rhodosporidium diobovatum Q95

Marine Drugs ◽  
2020 ◽  
Vol 18 (11) ◽  
pp. 546
Author(s):  
Jie Pan ◽  
Ni-Na Wang ◽  
Xue-Jing Yin ◽  
Xiao-Ling Liang ◽  
Zhi-Peng Wang
Keyword(s):  
Gallic Acid ◽  
Tannic Acid ◽  
Specific Activity ◽  
Maximum Activity ◽  
Sds Page ◽  
Rhodosporidium Diobovatum ◽  
Bacteria And Fungi ◽  
Ph Range ◽  
First Time

Tannase plays a crucial role in many fields, such as the pharmaceutical industry, beverage processing, and brewing. Although many tannases derived from bacteria and fungi have been thoroughly studied, those with good pH stabilities are still less reported. In this work, a mangrove-derived yeast strain Rhodosporidium diobovatum Q95, capable of efficiently degrading tannin, was screened to induce tannase, which exhibited an activity of up to 26.4 U/mL after 48 h cultivation in the presence of 15 g/L tannic acid. The tannase coding gene TANRD was cloned and expressed in Yarrowia lipolytica. The activity of recombinant tannase (named TanRd) was as high as 27.3 U/mL. TanRd was purified by chromatography and analysed by SDS-PAGE, showing a molecular weight of 75.1 kDa. The specific activity of TanRd towards tannic acid was 676.4 U/mg. Its highest activity was obtained at 40 °C, with more than 70% of the activity observed at 25–60 °C. Furthermore, it possessed at least 60% of the activity in a broad pH range of 2.5–6.5. Notably, TanRd was excellently stable at a pH range from 3.0 to 8.0; over 65% of its maximum activity remained after incubation. Besides, the broad substrate specificity of TanRd to esters of gallic acid has attracted wide attention. In view of the above, tannase resources were developed from mangrove-derived yeasts for the first time in this study. This tannase can become a promising material in tannin biodegradation and gallic acid production.

Coagulation performance of cucurbit[8]uril for the removal of azo dyes: effect of solution chemistry and coagulant dose

2018 ◽  
Vol 78 (2) ◽  
pp. 415-423 ◽  
Author(s):  
Wendong Wang ◽  
Zhiwen Chen ◽  
Kun Wu ◽  
Zongkuan Liu ◽  
Shengjiong Yang ◽  
...  
Keyword(s):  
Solution Chemistry ◽  
Color Removal ◽  
Dye Wastewater ◽  
Removal Rates ◽  
Molecular Weights ◽  
Coagulation Performance ◽  
Wide Ph Range ◽  
Ph Range ◽  
Significant Attention ◽  
Cr Removal

Abstract Dye wastewater has attracted significant attention because of its wide pH range and high content of color. In this work, the coagulation performances of cucurbit[8]uril for the removal of color from acid red 1 (AR1), orange II (OII), and Congo red (CR) dye wastewaters were investigated. Experimental results showed that color removal rates of greater than 95% for AR1, OII and CR were achieved at pH 6.0, when the dosage of cucurbit[8]uril was 1.51, 3.01 and 0.38 mmol·L−1, respectively. Under identical conditions, the color removal efficiencies of AR1 and CR were higher than OII, due to the larger molecular weights and more active hydroxyl and amino groups. Moreover, steady increases in AR1, OII and CR removal rates were recorded with increasing ionic strength. Such increases may be related to the reduction in thickness of the surface solvent membrane surrounding the dye colloids at high ionic strengths. Furthermore, Fourier transform infrared spectra demonstrated that no new bonds or functional groups were formed during coagulation, which indicates that the removal of AR1, OII and CR was primarily a physical process. The hydrogen bonds and inclusion complexes formed between cucurbit[8]uril and AR1, OII and CR contributed to the removal of color in coagulation predominantly.

Purification and characterization of catalase HPII from Escherichia coli K12

1986 ◽  
Vol 64 (7) ◽  
pp. 638-646 ◽  
Author(s):  
Peter C. Loewen ◽  
Jacek Switala
Keyword(s):  
Escherichia Coli ◽  
Sodium Dodecyl ◽  
Purification And Characterization ◽  
Unusual Structure ◽  
Escherichia Coli K12 ◽  
Molecular Weights ◽  
Ph Range ◽  
Heme Cofactor ◽  
Native Molecular Weight

Catalase (hydroperoxidase II or HPII) of Escherichia coli K12 has been purified using a protocol that also allows the purification of the second catalase HPI in large amounts. The purified HPII was found to have equal amounts of two subunits with molecular weights of 90 000 and 92 000. Only a single 92 000 subunit was present in the immunoprecipitate created when HPII antiserum was added directly to a crude extract, suggesting that proteolysis was responsible for the smaller subunit. The apparent native molecular weight was determined to be 532 000, suggesting a hexamer structure for the enzyme, an unusual structure for a catalase. HPII was very stable, remaining maximally active over the pH range 4–11 and retaining activity even in a solution of 0.1% sodium dodecyl sulfate and 7 M urea. The heme cofactor associated with HPII was also unusual for a catalase, in resembling heme d (a2) both spectrally and in terms of solubility. On the basis of heme-associated iron, six heme groups were associated with each molecule of enzyme or one per subunit.

Acid and alkaline phosphatases of Capnocytophaga species. II. Isolation, purification, and characterization of the enzymes from Capnocytophaga ochracea

1983 ◽  
Vol 29 (10) ◽  
pp. 1361-1368 ◽  
Author(s):  
Thomas P. Poirier ◽  
Stanley C. Holt
Keyword(s):  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Alkaline Phosphatases ◽  
Purification And Characterization ◽  
Molecular Weights ◽  
Sds Page ◽  
Kinetics Of

Capnocytophaga ochracea acid (AcP; EC 3.1.3.2) and alkaline (AlP; EC 3.1.3.1) phosphatase was isolated by Ribi cell disruption and purified by sodium dodecyl sulphate – polyacrylamide gel electrophoresis (SDS–PAGE.) Both phosphatases eluted from Sephadex G-150 consistent with molecular weights (migration) of 140 000 and 110 000. SDS–PAGE demonstrated a 72 000 and 55 000 subunit molecular migration for AcP and AlP, respectively. The kinetics of activity of purified AcP and AIP on p-nitrophenol phosphate and phosphoseryl residues of the phosphoproteins are presented.

scholarly journals Purification, Characterization of an Entomopathogenic Fungal Lectin from Purpureocillium Lilacinum, and its Involvement in Pathogenesis Leading to Mycotic Keratitis

2021 ◽  
Author(s):  
Narasimhappagari Jagadeesh ◽  
Supreeth Kulkarni ◽  
Vishwanath B Chachadi ◽  
Sanhita Roy ◽  
Shashikala Inamdar
Keyword(s):  
Proinflammatory Response ◽  
Corneal Epithelial ◽  
Purpureocillium Lilacinum ◽  
Ammonium Sulphate Precipitation ◽  
Reducing Conditions ◽  
Sds Page ◽  
Fungal Lectin ◽  
Human Corneal Epithelial Cells ◽  
Ph Range

Abstract A lectin PCL, from Purpureocillium lilacinum a saprophytic, filamentous fungus was purified from the crude extract of the mycelia using 70% ammonium sulphate precipitation followed by affinity chromatography on mucin-Sepharose 4 B column. PCL is a monomer with an apparent molecular mass of 18.5 kDa as revealed by SDS-PAGE under both reducing and non reducing conditions. PCL is a blood group non specific lectin and has highest affinity towards Chitin, Mucin, asialo mucin, Fetuin with a MIC of 0.15µg/mL and also recognizes L-fucose, galactose, lactose, N-acetly galactosamine, Hyaluronic acid. PCL is stable up to 60 ºC and within the pH range 4–8. To understand its role in pathogenesis, effect of PCL was evaluated on Human Corneal Epithelial Cells (HCECs). PCL showed strong glycan mediated binding to HCECsand PCL showed proinflammatory response at lower concentrations by stimulating secretion of IL-6, 8. In contrast PCL at higher concentrations revealed opposite effect of HCECs growth inhibition. All these results collectively support the involvement of PCL in mediating host pathogen interactions possibly leading to pathogenesis. In addition, considering the entomopathogenic effect of Purpureocillium lilacinum, PCL may be attributed for this beneficiary effect, which needs to be explored.

scholarly journals Tuning the Properties of High Molecular Weight Chitosans to Develop Full Water Solubility Within a Wide pH Range

2020 ◽  
Author(s):  
Anderson Fiamingo ◽  
Sergio Paulo Campana Filho ◽  
Osvaldo Novais Oliveira Junior
Keyword(s):  
Molecular Weight ◽  
Biomedical Applications ◽  
Random Distribution ◽  
Water Solubility ◽  
Aqueous Media ◽  
Degree Of Polymerization ◽  
Molecular Weights ◽  
H Nmr ◽  
Wide Ph Range ◽  
Ph Range

<p>The preparation of chitosans soluble in physiological conditions has been sought for years, but so far solubility in non-acidic aqueous media has only been achieved at the expense of lowering chitosan molecular weight. In this work, we applied the multistep ultrasound-assisted deacetylation process (USAD process) to β-chitin and obtained extensively deacetylated chitosans with high molecular weights (Mw ≥ 1,000,000 g mol<sup>-1</sup>). The homogeneous <i>N</i>-acetylation of a chitosan sample resulting from three consecutive USAD procedures allowed us to produce chitosans with a high weight average degree of polymerization (DPw ≈ 6,000) and tunable degrees of acetylation (DA from 5 to 80%). <i>N</i>-acetylation was carried out under mild conditions to minimize depolymerization, while preserving a predominantly random distribution of 2-amino-2-deoxy-D-glucopyanose (<i>GlcN</i>) and 2-acetamido-2-deoxy-D-glucopyanose (<i>GlcNAc</i>) units. This close to random distribution, inferred with deconvolution of nuclear magnetic resonance (<sup>1</sup>H NMR) spectra, is considered as responsible for the solubility within a wide pH range. Two of the highly <i>N</i>-acetylated chitosans (DA ≈ 60 % and ≈ 70 %) exhibited full water solubility even at neutral pH, which can expand the biomedical applications of chitosans. </p>

scholarly journals Biochemical Characterization of a New β-Agarase from Cellulophaga Algicola

2019 ◽  
Vol 20 (9) ◽  
pp. 2143 ◽  
Author(s):  
Han ◽  
Zhang ◽  
Yang
Keyword(s):  
Structural Model ◽  
Biochemical Characterization ◽  
Maximum Activity ◽  
Optimal Temperature ◽  
Salt Tolerant ◽  
Wide Ph Range ◽  
Specific Activities ◽  
Ph Range ◽  
Layer Chromatography

Cellulophaga algicola DSM 14237, isolated from the Eastern Antarctic coastal zone, was found to be able to hydrolyze several types of polysaccharide materials. In this study, a predicted β-agarase (CaAga1) from C. algicola was heterologously expressed in Escherichia coli. The purified recombinant CaAga1 showed specific activities of 29.39, 20.20, 14.12, and 8.99 U/mg toward agarose, pure agar, and crude agars from Gracilaria lemaneiformis and Porphyra haitanensis, respectively. CaAga1 exhibited an optimal temperature and pH of 40 oC and 7, respectively. CaAga1 was stable over a wide pH range from 4 to 11. The recombinant enzyme showed an unusual thermostability, that is, it was stable at temperature below or equal to 40oC and around 70 oC, but was thermolabile at about 50 oC. With the agarose as the substrate, the Km and Vmax values for CaAga1 were 1.19 mg/mL and 36.21 U/mg, respectively. The reducing reagent (dithiothreitol) enhanced the activity of CaAga1 by more than one fold. In addition, CaAga1 was salt-tolerant given that it retained approximately 70% of the maximum activity in the presence of 2 M NaCl. The thin layer chromatography results indicated that CaAga1 is an endo-type β-agarase and efficiently hydrolyzed agarose into neoagarotetraose (NA4) and neoagarohexaose (NA6). A structural model of CaAga1 in complex with neoagarooctaose (NA8) was built by homology modeling and explained the hydrolysis pattern of CaAga1.

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