Production of pectin lyase from Geobacillus pallidus p26, purification, characterization and fruit juice application

2014 ◽  
Vol 7 (1) ◽  
pp. 57-63 ◽  
Author(s):  
Safinur Yildirim Çelik ◽  
Nazan Demir ◽  
Yaşar Demir ◽  
Ahmet Adiguzel ◽  
Medine Gulluce
Keyword(s):  
Molecular Weight ◽  
Fruit Juice ◽  
Hot Spring ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Pectin Lyase ◽  
Anion Exchange Column ◽  
Sds Page ◽  
Temperature Optima ◽  
Geobacillus Pallidus

Abstract A bacterial strain was isolated from Pasinler hot spring, Erzurum, Turkey. The purified thermophilic isolate was identified as Geobacillus pallidus P26 and used to produce extracellular pectin lyase (EC 4.2.2.10). Pectin lyase enzyme was purified 34 fold by using DEAE-cellulose anion exchange column chromatography and characterized. Molecular weight of the enzyme was determined as 56 kDa by using Sephadex G-100 gel filtration chromatography. Purification of enzyme was verified by SDS-PAGE. The pH- and temperature optima of enzyme were determined (pH 9.0 and 60°C, respectively). Pectin lyase was mostly stable at 50 oC for 24 hours. Its’ activity decreased to 50 % for 24 h at 60°C. KM and Vmax were calculated as 24.8 mg/mL and 2.28 μmol/L min, respectively. Purified pectin lyase was inhibited by Fe3+, Zn2+, Cu2+, Ca2+, Co2+ and Hg2+ but not by Mg2+. The purified pectin lyase enzyme was used for getting fruits juices. It was found that yields of fruits juices increased when they were compared with control.

scholarly journals PREPARATION, PURIFICATION AND PROPERTIES OF LIPASE FROM HEPATOPANCREAS OF TRA (PANGASIUS) CATFISH

2011 ◽  
Vol 14 (3) ◽  
pp. 5-11
Author(s):  
Thy Bao Vuong ◽  
Lam Bich Tran ◽  
Duan Luu
Keyword(s):  
Heavy Metals ◽  
Molecular Weight ◽  
Enzyme Activity ◽  
Crude Extract ◽  
Gel Electrophoresis ◽  
Specific Activity ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Purification And Properties ◽  
Temperature Optima

Lipase from the hepatopancreas of Tra (Pangasius) catfish was purified by ammonium sulfate fractionation, followed by ion-exhange chromatography on DEAE Cellulose and gel filtration Sephadex G-75. The preparation was homogeneous on polyacrylamide disc gel electrophoresis. The specific activity of the purified enzyme was 37.95 times higher than that of the crude extract. The enzyme showed a molecular weight of 57000 Da. The pH and temperature optima of purified lipase were 8 and 500C respectively. Enzyme activity was enhanced by Ca2+ but inhibited by heavy metals Zn2+, Cd2+, Mg2+.

scholarly journals Isolasi dan Karakterisasi Inulinase dari Aspergillus niger Gmn11.1 Galur Lokal

2012 ◽  
Vol 11 (1) ◽  
pp. 19 ◽  
Author(s):  
Saryono Saryono
Keyword(s):  
Molecular Weight ◽  
Aspergillus Niger ◽  
Incubation Time ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Salt Precipitation ◽  
Sds Page ◽  
Optimum Ph ◽  
Relative Molecular Weight ◽  
Optimum Ph And Temperature

Inulin is a naturally potential polysaccharide used to produced fructose and fructooligosaccharide. Inulinaseknown also as ß-fructosidase can hydrolise inulin to fructose or fructooligosaccharide. Inulinase production fromAspergillus niger Gmn11.1 isolated from dahlia tubers is conducted using medium containing 1% inulin and 0,2%yeast extract. The crude enzyme (filtrate culture) is purified by means of ammonium sulphate salt precipitation,followed by Sephadex G25 gel filtration column chromatography and DEAE cellulose anion exchanger columnchromatography. The result indicated that the enzyme had optimum pH and temperature of 4,6 and 450C, respectivelywith incubation time of 15 hours. The Km and Vmaxs values obtained from this experiment are 20 mg/ml and 0,769mg/ml/hours, respectively. Whereas the relative molecular weight of inulinase was monitored by SDS PAGE is 63KDa.

scholarly journals Purification and Characterization of Glucose-6-Phosphate Dehydrogenase from Camel Liver

2014 ◽  
Vol 2014 ◽  
pp. 1-10 ◽  
Author(s):  
Mahmoud A. Ibrahim ◽  
Abdel-Hady M. Ghazy ◽  
Ahmed M. H. Salem ◽  
Mohamed A. Ghazy ◽  
Mohamed M. Abdel-Monsef
Keyword(s):  
Molecular Weight ◽  
Specific Activity ◽  
Divalent Cations ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Native Form ◽  
Native Page ◽  
Sds Page ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Sepharose 4B

Glucose-6-phosphate dehydrogenase from camel liver was purified to homogeneity by ammonium sulfate precipitation and a combination of DEAE-cellulose, Sephacryl S-300 gel filtration, and 2′, 5′ ADP Sepharose 4B affinity chromatography columns. The specific activity of camel liver G6PD is increased to 1.80438 units/mg proteins with 63-fold purification. It turned out to be homogenous on both native PAGE and 12% SDS PAGE, with a molecular weight of 64 kDa. The molecular weight of the native form of camel liver G6PD was determined to be 194 kDa by gel filtration indicating a trimeric protein. The Km value was found to be 0.081 mM of NADP+. Camel liver G6PD displayed its optimum activity at pH 7.8 with an isoelectric point (pI) of pH 6.6–6.8. The divalent cations MgCl2, MnCl2, and CoCl2 act as activators; on the other hand, CaCl2 and NiCl2 act as moderate inhibitors, while FeCl2, CuCl2, and ZnCl2 are potent inhibitors of camel liver G6PD activity. NADPH inhibited camel liver G6PD competitively with Ki value of 0.035 mM. One binding site was deduced for NADPH on the enzyme molecule. This study presents a simple and reproducible purification procedure of G6PD from the camel liver.

Characterization of an isozyme of S-adenosylmethionine synthetase from rat liver

1984 ◽  
Vol 62 (5) ◽  
pp. 276-279 ◽  
Author(s):  
C. H. Lin ◽  
W. Chung ◽  
K. P. Strickland ◽  
A. J. Hudson
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Amino Acid ◽  
Enzymatic Synthesis ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Aromatic Amino Acids ◽  
Adenosylmethionine Synthetase ◽  
Cellulose Column

An isozyme of S-adenosylmethionine synthetase has been purified to homogeneity by ammonium sulfate fractionation, DEAE-cellulose column chromatography, and gel filtration on a Sephadex G-200 column. The purified enzyme is very unstable and has a molecular weight of 120 000 consisting of two identical subunits. Amino acid analysis on the purified enzyme showed glycine, glutamate, and aspartate to be the most abundant and the aromatic amino acids to be the least abundant. It possesses tripolyphosphatase activity which can be stimulated five to six times by S-adenosylmethionine (20–40 μM). The findings support the conclusion that an enzyme-bound tripolyphosphate is an obligatory intermediate in the enzymatic synthesis of S-adenosylmethionine from ATP and methionine.

scholarly journals A chromatographic preparation of ox growth hormone

1966 ◽  
Vol 100 (3) ◽  
pp. 593-600 ◽  
Author(s):  
M Wallis ◽  
HBF Dixon
Keyword(s):  
Molecular Weight ◽  
Growth Hormone ◽  
Gel Electrophoresis ◽  
Anterior Pituitary ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Starch Gel Electrophoresis ◽  
Highly Active ◽  
Cross Linkage ◽  
Starch Gel

1. A method is described for the chromatographic preparation of ox growth hormone. It involves chromatography of an extract of anterior pituitary lobes on DEAE-cellulose, followed by rechromatography on a dextran gel of low cross-linkage (Sephadex G-100). 2. The product is highly active in growth-hormone assays, and is obtained in good yield. It was homogeneous by several criteria, but showed some heterogeneity on starch-gel electrophoresis. 3. The molecular weight of the hormone was estimated from its behaviour on gel-filtration columns under various conditions. Evidence that the hormone may dissociate into sub-units under some conditions is presented.

scholarly journals The isocitrate dehydrogenases of Acinetobacter lwoffi. Separation and properties of two nicotinamide–adenine dinucleotide phosphate-linked isoenzymes

1972 ◽  
Vol 130 (1) ◽  
pp. 211-219 ◽  
Author(s):  
Colin H. Self ◽  
P. David J. Weitzman
Keyword(s):  
Molecular Weight ◽  
Ph Dependence ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Molecular Size ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Low Concentrations ◽  
Stimulation Of

Two isoenzymes of NADP-linked isocitrate dehydrogenase have been identified in Acinetobacter lwoffi and have been termed isoenzyme-I and isoenzyme-II. The isoenzymes may be separated by ion-exchange chromatography on DEAE-cellulose, by gel filtration on Sephadex G-200, or by zonal ultracentrifugation in a sucrose gradient. Low concentrations of glyoxylate or pyruvate effect considerable stimulation of the activity of isoenzyme-II. The isoenzymes also differ in pH-dependence of activity, kinetic parameters, stability to heat or urea and molecular size. Whereas isoenzyme-I resembles the NADP-linked isocitrate dehydrogenases from other organisms in having a molecular weight under 100000, isoenzyme-II is a much larger enzyme (molecular weight around 300000) resembling the NAD-linked isocitrate dehydrogenases of higher organisms.

scholarly journals α-Crystallin. The isolation and characterization of distinct macromolecular fractions

1971 ◽  
Vol 124 (2) ◽  
pp. 337-343 ◽  
Author(s):  
Abraham Spector ◽  
Lu-Ku Li ◽  
Robert C. Augusteyn ◽  
Arthur Schneider ◽  
Thomas Freund
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Chemical Difference ◽  
Molecular Weights ◽  
Isolation And Characterization ◽  
Different Populations ◽  
Molecular Weight Fractions

α-Crystallin was isolated from calf lens periphery by chromatography on DEAE-cellulose and gel filtration. Three distinct populations of macromolecules have been isolated with molecular weights in the ranges approx. 6×105−9×105, 0.9×106−4×106and greater than 10×106. The concentration of macromolecules at the molecular-weight limits of a population are very low. The members of the different populations do not appear to be in equilibrium with each other. Further, in those molecular-weight fractions investigated, no equilibrium between members of the same population was observed. The population of lowest molecular weight comprises 65–75% of the total material. The amino acid and subunit composition of the different-sized fractions appear very similar, if not identical. The only chemical difference observed between the fractions is the presence of significant amounts of sugar in the higher-molecular-weight fractions. Subunit molecular weights of approx. 19.5×103and 22.5×103were observed for all α-crystallin fractions.

scholarly journals Isolation and Characterization of Two Proteins fromMoraxella catarrhalis That Bear a Common Epitope

1998 ◽  
Vol 66 (9) ◽  
pp. 4374-4381 ◽  
Author(s):  
John C. McMichael ◽  
Michael J. Fiske ◽  
Ross A. Fredenburg ◽  
Deb N. Chakravarti ◽  
Karl R. VanDerMeid ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Bacterial Attachment ◽  
Vaccine Candidates ◽  
Isolation And Characterization ◽  
Sds Page

ABSTRACT The UspA1 and UspA2 proteins of Moraxella catarrhalisare potential vaccine candidates for preventing disease caused by this organism. We have characterized both proteins and evaluated their vaccine potential using both in vitro and in vivo assays. Both proteins were purified from the O35E isolate by Triton X-100 extraction, followed by ion-exchange and hydroxyapatite chromatography. Analysis of the sequences of internal peptides, prepared by enzymatic and chemical cleavage of the proteins, revealed that UspA1 and UspA2 exhibited distinct structural differences but shared a common sequence including an epitope recognized by the monoclonal antibody 17C7. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), purified UspA1 exhibited a molecular weight of approximately 350,000 when unheated and a molecular weight of 100,000 after being heated for 10 min at 100°C. In contrast, purified UspA2 exhibited an apparent molecular weight of 240,000 by SDS-PAGE that did not change with the length of time of heating. Their sizes as determined by gel filtration were 1,150,000 and 830,000 for UspA1 and UspA2, respectively. Preliminary results indicate the proteins have separate functions in bacterial pathogenesis. Purified UspA1 was found to bind HEp-2 cells, and sera against UspA1, but not against UspA2, blocked binding of the O35E isolate to the HEp-2 cells. UspA1 also bound fibronectin and appears to have a role in bacterial attachment. Purified UspA2, however, did not bind fibronectin but had an affinity for vitronectin. Both proteins elicited bactericidal antibodies in mice to homologous and heterologous disease isolates. Finally, mice immunized with each of the proteins, followed by pulmonary challenge with either the homologous or a heterologous isolate, cleared the bacteria more rapidly than mock-immunized mice. These results suggest that UspA1 and UspA2 serve different virulence functions and that both are promising vaccine candidates.

scholarly journals PURIFICATION AND PARTIAL CHARACTERIZATION OF L-ASPARAGINASE ENZYME PRODUCED BY NEWLY ISOLATE BACILLUS SP.

2017 ◽  
Vol 18 (2) ◽  
pp. 1-10 ◽  
Author(s):  
Dzun Noraini Jimat ◽  
Intan Baizura Firda Mohamed ◽  
Azlin Suhaida Azmi ◽  
Parveen Jamal
Keyword(s):  
Specific Activity ◽  
Hot Spring ◽  
Gel Filtration ◽  
Maximum Activity ◽  
Exchange Chromatography ◽  
Bacillus Sp ◽  
Sds Page ◽  
Fast Flow ◽  
Microbial Strain

A newly bacterial producing L-asparaginase was successful isolated from Sungai Klah Hot Spring, Perak, Malaysia and identified as Bacillus sp. It was the best L-asparaginase producer as compared to other isolates. Production of L-asparaginase from the microbial strain was carried out under liquid fermentation. The crude enzyme was then centrifuged and precipitated with ammonium sulfate before further purified with chromatographic method. The ion exchange chromatography HiTrap DEAE-Sepharose Fast Flow column followed by separation on Superose 12 gel filtration were used to obtain pure enzyme. The purified enzyme showed 10.11 U/mg of specific activity, 50.07% yield with 2.21 fold purification. The purified enzyme was found to be dimer in form, with a molecular weight of 65 kDa as estimated by SDS-PAGE. The maximum activity of the purified L-asparaginase was observed at pH 9 and temperature of 60°C.

scholarly journals Coal Depolymerising Activity and Haloperoxidase Activity of Mn Peroxidase fromFomes durissimusMTCC-1173

2011 ◽  
Vol 2011 ◽  
pp. 1-8 ◽  
Author(s):  
Sunil Kumar Singh ◽  
Meera Yadav ◽  
Sudha Yadava ◽  
Kapil Deo Singh Yadav
Keyword(s):  
Deae Cellulose ◽  
Anion Exchange Chromatography ◽  
Sodium Lactate ◽  
Low Ph ◽  
Exchange Chromatography ◽  
Fungal Strain ◽  
Mn Peroxidase ◽  
Sds Page ◽  
Catalytic Rate ◽  
Temperature Optima

Mn peroxidase has been purified to homogeneity from the culture filtrate of a new fungal strainFomes durissimusMTCC-1173 using concentration by ultrafiltration and anion exchange chromatography on diethylaminoethyl (DEAE) cellulose. The molecular mass of the purified enzyme has been found to be 42.0 kDa using SDS-PAGE analysis. The values using MnSO4and H2O2as the variable substrates in 50 mM lactic acid-sodium lactate buffer pH 4.5 at were 59 μM and 32 μM, respectively. The catalytic rate constants using MnSO4and H2O2were 22.4 s−1and 14.0 s−1, respectively, giving the values of 0.38 μM−1s−1and 0.44 μM−1s−1, respectively. The pH and temperature optima of the Mn peroxidase were 4 and , respectively. The purified MnP depolymerises humic acid in presence of H2O2. The purified Mn peroxidase exhibits haloperoxidase activity at low pH.

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