scholarly journals Production, Purification, and Characterization of a MajorPenicillium glabrumXylanase Using Brewer's Spent Grain as Substrate

2013 ◽  
Vol 2013 ◽  
pp. 1-8 ◽  
Author(s):  
Adriana Knob ◽  
Susan Michelz Beitel ◽  
Diana Fortkamp ◽  
César Rafael Fanchini Terrasan ◽  
Alex Fernando de Almeida
Keyword(s):  
Xylanase Activity ◽  
Stationary Condition ◽  
Purification And Characterization ◽  
Xylanase Production ◽  
Brewer’S Spent Grain ◽  
Agroindustrial Waste ◽  
Sds Page ◽  
Exclusion Chromatography ◽  
Spent Grain

In recent decades, xylanases have been used in many processing industries. This study describes the xylanase production byPenicillium glabrumusing brewer's spent grain as substrate. Additionally, this is the first work that reports the purification and characterization of a xylanase using this agroindustrial waste. Optimal production was obtained whenP. glabrumwas grown in liquid medium in pH 5.5, at 25 °C, under stationary condition for six days. The xylanase fromP. glabrumwas purified to homogeneity by a rapid and inexpensive procedure, using ammonium sulfate fractionation and molecular exclusion chromatography. SDS-PAGE analysis revealed one band with estimated molecular mass of 18.36 kDa. The optimum activity was observed at 60 °C, in pH 3.0. The enzyme was very stable at 50 °C, and high pH stability was verified from pH 2.5 to 5.0. The ion Mn2+and the reducing agentsβ-mercaptoethanol and DTT enhanced xylanase activity, while the ions Hg2+, Zn2+, and Cu2+as well as the detergent SDS were strong inhibitors of the enzyme. The use of brewer's spent grain as substrate for xylanase production cannot only add value and decrease the amount of this waste but also reduce the xylanase production cost.

Purification and characterization of an exochitinase from Bacillus thuringiensis subsp. aizawai and its action against phytopathogenic fungi

2006 ◽  
Vol 52 (7) ◽  
pp. 651-657 ◽  
Author(s):  
Luis Morales de la Vega ◽  
J Eleazar Barboza-Corona ◽  
Maria G Aguilar-Uscanga ◽  
Mario Ramírez-Lepe
Keyword(s):  
Bacillus Thuringiensis ◽  
Sodium Dodecyl ◽  
Sclerotium Rolfsii ◽  
Phytopathogenic Fungi ◽  
Purification And Characterization ◽  
Chitinolytic Enzyme ◽  
Bacillus Thuringiensis Subsp ◽  
Sds Page ◽  
Fusarium Sp

A chitinolytic enzyme from Bacillus thuringiensis subsp. aizawai has been purified and its molecular mass was estimated ca. 66 kDa by sodium dodecyl sulfate – polyacryamide gel electrophoresis (SDS–PAGE). The enzyme was able to hydrolyze chitin to chitobiosides but not carboxymethylcellulose, cellulose, pullulan, and laminarin. Optimal pH and temperature were detected at 6 and 50 °C, respectively. Stability, in the absence of substrate, was observed at temperatures less than 60 °C and pH between 5 and 8. Enzyme activity was significantly inhibited by K+ and EDTA and completely inhibited by Hg2+. Purified chitinase showed lytic activity against cell walls from six phytopathogenic fungi and inhibited the mycelial growth of both Fusarium sp. and Sclerotium rolfsii. The biocontrol efficacy of the enzyme was tested in the protection of bean seeds infested with six phytopathogenic fungi.Key words: chitinase, Bacillus thuringiensis, purification, phytopathogenic fungi.

scholarly journals Partial purification and characterization of xylanase from Bacillus cereus X3

2014 ◽  
Vol 11 (2) ◽  
pp. 1056-1061
Author(s):  
Baghdad Science Journal
Keyword(s):  
Bacillus Cereus ◽  
Maximal Activity ◽  
Partial Purification ◽  
Purification And Characterization ◽  
Xylanase Production ◽  
Optimum Activity ◽  
Biology Department ◽  
Characterization Study ◽  
Quantitative Screening

Three strain of Bacillus cereus were obtained from soil sours Laboratories of Biology Department/ College of Science/ University of Baghdad. The bacteria secreted extracellular xylanase in liquid cultur the test ability of xylanase production from these isolates was studied semi quantitative and quantitative screening appeared that Bacillus cereus X3 was the highest xylanase producer. The enzyme was partial purification 191 fold from cultur by reached step by 4 U/mg proteins by ammonium sulfat precipitation 80%, Ion exchang DEAE-cellulos chromatography Characterization study of the partial purifation enzyme revealed that the enzyme had a optimum activity pH8 and activity was stable in the pH rang (8-10) for 30min. maximal activity was attained at 50C

Cloning and Characterization of a Novel N-acetylglucosaminidase (AtlD) from Enterococcus faecalis

2018 ◽  
Vol 28 (1) ◽  
pp. 14-27 ◽  
Author(s):  
Carlos Eduardo Serrano-Maldonado ◽  
Israel García-Cano ◽  
Augusto González-Canto ◽  
Eliel Ruiz-May ◽  
Jose Miguel Elizalde-Contreras ◽  
...  
Keyword(s):  
Enterococcus Faecalis ◽  
Food Industry ◽  
Pathogenic Bacteria ◽  
Heterologous Protein ◽  
Biochemical Characterization ◽  
Ph Values ◽  
Food Borne ◽  
Sds Page ◽  
Exclusion Chromatography

The <i>atlD</i> gene from<i></i> an <i>Enterococcus faecalis</i> strain isolated from a Mexican artisanal cheese was cloned, sequenced and expressed in <i>Escherichia coli</i> in order to perform a biochemical characterization<i>.</i> A partial amino acid sequence of the heterologous protein was obtained by LC-MS/MS, and it corresponded to a novel peptidoglycan hydrolase designated AtlD. Its molecular mass was 62–75 kDa, as determined by SDS-PAGE, zymography, Western blot, and exclusion chromatography. Electrofocusing rendered an isoelectric point (pI) of 4.8. It exhibited N-acetylglucosaminidase activity, with an optimal pH and temperature between 6–7 and 50°C, respectively. It retained 85% activity with NaCl at 1,000 mM, but it was susceptible to divalent ions, particularly Zn<sup>2+</sup>. It showed antibacterial activity against <i>Listeria monocytogenes</i>,<i> Staphylococcus aureus</i>, and enterococcal<i></i> strains of clinical origin. Due to the fact that it showed activity versus pathogenic bacteria, and because of its capabilities under ionic strength, temperature, and pH values present in food matrices, it could be applied as an additive in the food industry. This study will aid in the design of new antibacterial agents of natural origin to combat food-borne diseases, and it could be used as an industrial or hospital hygiene agent as well.

Purification and Characterization of Cytochrome c6 from the Unicellular Green Alga Scenedesmus obliquus

1997 ◽  
Vol 52 (11-12) ◽  
pp. 740-746 ◽  
Author(s):  
Röbbe Wünschiers ◽  
Thomas Zinn ◽  
Dietmar Linder ◽  
Rüdiger Schulz
Keyword(s):  
Cytochrome C ◽  
Green Alga ◽  
Scenedesmus Obliquus ◽  
Terminal Sequence ◽  
Ammonium Sulfate Precipitation ◽  
Purification And Characterization ◽  
Unicellular Green Alga ◽  
Sds Page ◽  
Monoraphidium Braunii

Abstract Purification of a soluble cytochrome c6 from the unicellular green alga Scenedesmus obliquus by a simple and rapid method is described. The purification procedure includes ammonium sulfate precipitation and non-denaturating PAGE. The N-terminal sequence of the first 20 amino acids was determined and shows 85% similarity and 75% identity to the sequence of cytochrome c6 from the green alga Monoraphidium braunii. The ferrocyto-chrome shows typical UV/VIS absorption peaks at 552.9, 521.9 and 415.7 nm. The apparent molecular mass was estimated to be 12 kD a by SDS-PAGE. EPR-spectroscopy at 20K shows resonances indicative for two distinct low-spin heme forms.

scholarly journals Purification and characterization of pig kidney aminopeptidase P. A glycosyl-phosphatidylinositol-anchored ectoenzyme

1990 ◽  
Vol 267 (2) ◽  
pp. 509-515 ◽  
Author(s):  
N M Hooper ◽  
J Hryszko ◽  
A J Turner
Keyword(s):  
Chelating Agents ◽  
Inhibitory Effect ◽  
Purification And Characterization ◽  
Glycosyl Phosphatidylinositol ◽  
Pig Kidney ◽  
Sds Page ◽  
Cyclic Phosphate ◽  
Aminopeptidase P ◽  
Mild Acid

Aminopeptidase P (EC 3.4.11.9) was solubilized from pig kidney membranes with bacterial phosphatidylinositol-specific phospholipase C (PI-PLC) and then purified by a combination of anion-exchange and hydrophobic-interaction chromatographies. Contaminating peptidase activities were removed by selective affinity chromatography. The purified enzyme was apparently homogeneous on SDS/PAGE with an Mr of 91,000. Enzymic deglycosylation revealed that aminopeptidase P is a glycoprotein, with up to 25% by weight of the protein being due to the presence of N-linked sugars. The phospholipase-solubilized aminopeptidase P was recognized by an antiserum to the cross-reacting determinant (CRD) characteristic of the glycosyl-phosphatidylinositol anchor. This recognition was abolished by mild acid treatment or deamination with HNO2, indicating that the CRD was due exclusively to the inositol 1,2-cyclic phosphate ring epitope generated by the action of PI-PLC. The activity of aminopeptidase P was inhibited by chelating agents and was stimulated by Mn2+ or Co2+ ions, confirming the metallo-enzyme nature of this peptidase. Selective inhibitors of other aminopeptidases (actinonin, amastatin, bestatin and puromycin) had little or no inhibitory effect.

scholarly journals Molecular cloning and biochemical characterization of rabbit factor XI

2002 ◽  
Vol 367 (1) ◽  
pp. 49-56 ◽  
Author(s):  
Dipali SINHA ◽  
Mariola MARCINKIEWICZ ◽  
David GAILANI ◽  
Peter N. WALSH
Keyword(s):  
Human Factor ◽  
Biochemical Characterization ◽  
Protein A ◽  
Size Exclusion ◽  
Factor Xi ◽  
Sds Page ◽  
Exclusion Chromatography ◽  
Intracellular Process ◽  
And Function

Human factor XI, a plasma glycoprotein required for normal haemostasis, is a homodimer (160kDa) formed by a single interchain disulphide bond linking the Cys-321 of each Apple 4 domain. Bovine, porcine and murine factor XI are also disulphide-linked homodimers. Rabbit factor XI, however, is an 80kDa polypeptide on non-reducing SDS/PAGE, suggesting that rabbit factor XI exists and functions physiologically either as a monomer, as does prekallikrein, a structural homologue to factor XI, or as a non-covalent homodimer. We have investigated the structure and function of rabbit factor XI to gain insight into the relation between homodimeric structure and factor XI function. Characterization of the cDNA sequence of rabbit factor XI and its amino acid translation revealed that in the rabbit protein a His residue replaces the Cys-321 that forms the interchain disulphide linkage in human factor XI, explaining why rabbit factor XI is a monomer in non-reducing SDS/PAGE. On size-exclusion chromatography, however, purified plasma rabbit factor XI, like the human protein and unlike prekallikrein, eluted as a dimer, demonstrating that rabbit factor XI circulates as a non-covalent dimer. In functional assays rabbit factor XI and human factor XI behaved similarly. Both monomeric and dimeric factor XI were detected in extracts of cells expressing rabbit factor XI. We conclude that the failure of rabbit factor XI to form a covalent homodimer due to the replacement of Cys-321 with His does not impair its functional activity because it exists in plasma as a non-covalent homodimer and homodimerization is an intracellular process.

scholarly journals Purification and characterization of N-ethylmaleimide-insensitive phosphatidic acid phosphohydrolase (PAP2) from rat liver

1995 ◽  
Vol 308 (3) ◽  
pp. 983-989 ◽  
Author(s):  
I N Fleming ◽  
S J Yeaman
Keyword(s):  
Phosphatidic Acid ◽  
Rat Liver ◽  
Lysophosphatidic Acid ◽  
Gel Filtration ◽  
Purification And Characterization ◽  
Sds Page ◽  
Chromatography Columns ◽  
Triton X ◽  
Second Peak

N-Ethylmaleimide-insensitive phosphatidic acid phosphohydrolase (PAP; EC 3.1.3.4) was purified 5900-fold from rat liver. The enzyme was solubilized from membranes with octylglucoside, fractionated with (NH4)2SO4, and purified in the presence of Triton X-100 by chromatography on Sephacryl S300, hydroxyapatite, heparin-Sepharose and Affi-Gel Blue. Silver-stained SDS/PAGE indicated that the enzyme was an 83 kDa polypeptide. Sephacryl S-300 gel filtration also produced a second peak of enzyme activity, which was eluted from all of the chromatography columns at a different position from the purified enzyme. SDS/PAGE indicated that it contained three polypeptides (83 kDa, 54 kDa and 34 kDa), and gel filtration suggested that it was not an aggregate of the purified enzyme. Both forms were sensitive to inhibition by amphiphilic amines, Mn2+ and Zn2+, but not by N-ethylmaleimide. Purified PAP required detergent for activity, but was not activated by Mg2+, fatty acids or phospholipids. The enzyme was able to dephosphorylate lysophosphatidic acid or phosphatidic acid, and was inhibited by diacylglycerol and monoacylglycerol. No evidence was obtained for regulation of PAP by reversible phosphorylation.

Preparation and characterization of brewer’s spent grain protein-chitosan composite films

2015 ◽  
Vol 52 (11) ◽  
pp. 7549-7555 ◽  
Author(s):  
Ji-Hyeon Lee ◽  
Ji-Hyun Lee ◽  
Hyun-Ju Yang ◽  
Kyung Bin Song
Keyword(s):  
Composite Films ◽  
Grain Protein ◽  
Brewer’S Spent Grain ◽  
Spent Grain ◽  
Chitosan Composite

scholarly journals Production and Partial Characterization of an Alkaline Xylanase from a Novel FungusCladosporium oxysporum

2016 ◽  
Vol 2016 ◽  
pp. 1-7 ◽  
Author(s):  
Guo-Qiang Guan ◽  
Peng-Xiang Zhao ◽  
Jin Zhao ◽  
Mei-Juan Wang ◽  
Shu-Hao Huo ◽  
...  
Keyword(s):  
Nitrogen Source ◽  
Wheat Bran ◽  
Internal Transcribed Spacer ◽  
Gene Sequence ◽  
Agricultural Waste ◽  
Xylanase Activity ◽  
Maximum Activity ◽  
Alkaline Proteases ◽  
Xylanase Production

A new fungusCladosporium oxysporumGQ-3 producing extracellular xylanase was isolated from decaying agricultural waste and identified based on the morphology and comparison of internal transcribed spacer (ITS) rDNA gene sequence.C. oxysporumproduced maximum xylanase activity of 55.92 U/mL with wheat bran as a substrate and NH4Cl as a nitrogen source. Mg2+improvedC. oxysporumxylanase production.Partially purified xylanase exhibited maximum activity at 50°C and pH 8.0, respectively, and showed the stable activity after 2-h treatment in pH 7.0–8.5 or below 55°C. Mg2+enhanced the xylanase activity by 2% while Cu2+had the highest inhibition ratio of 57.9%. Furthermore,C. oxysporumxylanase was resistant to most of tested neutral and alkaline proteases. Our findings indicated thatCladosporium oxysporumGQ-3 was a novel xylanase producer, which could be used in the textile processes or paper/feed industries.

Important role of fungal intracellular laccase for melanin synthesis: purification and characterization of an intracellular laccase from Lentinula edodes fruit bodies

Microbiology ◽  
2003 ◽  
Vol 149 (9) ◽  
pp. 2455-2462 ◽  
Author(s):  
Masaru Nagai ◽  
Maki Kawata ◽  
Hisayuki Watanabe ◽  
Machiko Ogawa ◽  
Kumiko Saito ◽  
...  
Keyword(s):  
Lentinula Edodes ◽  
Veratryl Alcohol ◽  
Size Exclusion ◽  
Melanin Synthesis ◽  
Sds Page ◽  
Diammonium Salt ◽  
Exclusion Chromatography ◽  
Homogeneous Preparation

A laccase (EC 1.10.3.2) was isolated from the fully browned gills of Lentinula edodes fruit bodies. The enzyme was purified to a homogeneous preparation using hydrophobic, cation-exchange and size-exclusion chromatography. SDS-PAGE analysis showed the purified laccase, Lcc 2, to be a monomeric protein of 58·0 kDa. The enzyme had an isoelectric point of around pH 6·9. The optimum pH for enzyme activity was around 3·0 against 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt (ABTS), and it was most active at 40 °C and stable up to 50 °C. The enzyme contained 8·6 % carbohydrate and some copper atoms. The enzyme oxidized ABTS, p-phenylenediamine, pyrogallol, guaiacol, 2,6-dimethoxyphenol, catechol and ferulic acid, but not veratryl alcohol and tyrosine. β-(3,4-Dihydroxyphenyl)alanine (l-DOPA), which was not oxidized by a laccase previously reported from the culture filtrate of L. edodes, was also oxidized by Lcc 2, and the oxidative product of l-dopa was identified as l-DOPA quinone by HPLC analysis. Lcc 2 was able to oxidize phenolic compounds extracted from fresh gills to brown-coloured products, suggesting a role for laccase in melanin synthesis in this strain.

Sign in / Sign up

Export Citation Format

Share Document