scholarly journals Studies on mammalian glucoamylases with special reference to monkey intestinal glucoamylase

1970 ◽  
Vol 117 (5) ◽  
pp. 939-946 ◽  
Author(s):  
B. Seetharam ◽  
N. Swaminathan ◽  
A. N. Radhakrishnan
Keyword(s):  
Special Reference ◽  
Enrichment Factor ◽  
Intestinal Mucosa ◽  
Liver Enzymes ◽  
Gel Filtration ◽  
Maximum Activity ◽  
Similar Magnitude ◽  
Glucoamylase Activity ◽  
Specific Activities ◽  
Intestinal Enzyme

1. Highly purified preparations of glucoamylase were obtained from liver, spleen and intestine of the monkey. The enrichment factor was lower for intestine (60-fold) compared with that of liver (1200-fold) and of spleen (2000-fold) but the final specific activities were of a similar magnitude. 2. The liver and spleen enzymes had maximum activity at pH4.8 whereas the intestinal enzyme showed an optimum at pH5.8. The Km values for both starch and maltose with spleen and liver enzymes were higher than for the intestinal enzyme. With the intestinal enzyme, the Vmax. values were higher for both starch and maltose than those of the spleen and liver enzymes. 3. Gel filtration on Sephadex G-200 under identical conditions revealed that liver and spleen enzymes emerge from the columns much later than the intestinal enzyme. 4. Evidence is presented that the glucoamylase activity of the intestinal mucosa is exhibited by the maltase II fraction. 5. Tris, pentaerythritol and turanose inhibited glucoamylase from all the three tissues, but turanose inhibited the spleen and liver enzymes to a higher degree than the intestinal enzyme.

scholarly journals LYSOSOMAL ENZYMES OF RAT INTESTINAL MUCOSA

1964 ◽  
Vol 23 (2) ◽  
pp. 233-240 ◽  
Author(s):  
Lichu Hsu ◽  
A. L. Tappel
Keyword(s):  
Rat Liver ◽  
Intestinal Mucosa ◽  
Digestive Enzymes ◽  
Lysosomal Enzymes ◽  
Liver Enzymes ◽  
Enzyme Release ◽  
Centrifugal Forces ◽  
Physiological Functions ◽  
Specific Activities

Six intracellular hydrolases known to be associated with lysosomes in rat liver were found in rat intestinal mucosa. The extent to which they were particulate-bound and the degree of enzyme release when the particulate fractions were suspended in hypotonic media followed the same pattern in both mucosa and liver. The specific activities of the mucosa enzymes were either comparable to or slightly smaller than those of the liver enzymes. These results suggest that the mucosa hydrolases belong to lysosome-like particles. However, differential fractionation of the mucosa indicated that the particles from the mucosa sediment at lower centrifugal forces than do those from the liver and are more heterogeneous in size, bearing a closer resemblance to kidney lysosomes. Possible physiological functions of particulate-bound digestive enzymes in intestinal mucosa are discussed.

scholarly journals Production and optimization of Aspergillus niger glucoamylase using amylopectin from guinea corn starch as the sole carbon source

2017 ◽  
Vol 52 (4) ◽  
pp. 263-272
Author(s):  
PC Okwuenu ◽  
AL Ezugwu ◽  
FC Chilaka
Keyword(s):  
Carbon Source ◽  
Ammonium Sulphate ◽  
Sole Carbon Source ◽  
Corn Starch ◽  
Gel Filtration ◽  
Ph Optimum ◽  
Glucoamylase Activity ◽  
Ammonium Sulphate Precipitation ◽  
Specific Activities ◽  
Two Peaks

A Fourteen day experimental study was carried out to determine the day of highest glucoamylase activity using amylopectin from guinea corn starch as the sole carbon source. Two peaks of high activity were observed on the fifth and twelveth days, and were thus mass produced. Specific activities for crude enzymes were found to be 729.45 U/mg and 1046.82 U/mg for day five and twelve harvested enzymes respectively. Ammonium sulphate saturations, 70% and 20%, were found suitable to precipitate proteins with highest glucoamylase activity for day five and twelve harvested enzymes respectively. After ammonium sulphate precipitation and gel filtration, specific activities were found to be 65.98 U/mg and 180.52 U/mg respectively for day five harvested enzyme and 61.51 U/mg and 272.81 U/mg for day twelve harvested enzyme. The pH optimum for day five harvested enzyme were found to be 7.5, 7.5 and 6.0 using tiger nut, cassava and guinea corn starches as substrates respectively, also, the pH optimum for day twelve harvested enzyme were found to be 5.0, 8.5 and 7.0 using tiger nut, cassava and guinea corn starches as substrate, respectively. Optimum temperatures were found to be 50˚C and 45˚C for day five and twelve harvested enzymes, respectively. Km and Vmax, of day five harvested enzyme were found to be 770.75 mg/ml and 2500 μmol/min, 158.55 mg/ml and 500 μmol/min and 46.23 mg/ml and 454.53 μmol/min using cassava, guinea corn and tiger nut starches as substrate respectively. Km and Vmax of day twelve harvested enzyme were found to be 87.1 mg/ml and 384.61 μmol/min, 29.51 mg/ml and 243.90 μmol/min, and 2364 mg/ml and 2500 μmol/min, using cassava, guinea corn and tiger nut starches as substrate respectively.Bangladesh J. Sci. Ind. Res. 52(4), 263-272, 2017

scholarly journals Purification and characterization of acid phosphatase from a germinating black gram (Vigna mungo L.) seedling

2011 ◽  
Vol 63 (3) ◽  
pp. 747-756 ◽  
Author(s):  
A.K.M. Asaduzzaman ◽  
Habibur Rahman ◽  
Tanzima Yeasmin
Keyword(s):  
Acid Phosphatase ◽  
Gel Electrophoresis ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Inhibitory Effect ◽  
Maximum Activity ◽  
Exchange Chromatography ◽  
Black Gram ◽  
Km Value

An acid phosphatase has been isolated and purified from an extract of a germinating black gram seedling. The method was accomplished by gel filtration of a germinating black gram seedling crude extract on sephadex G-75 followed by ion exchange chromatography on DEAE cellulose. The acid phosphatase gave a single band on SDS-polyacrylamide slab gel electrophoresis. The molecular weight of the acid phosphatase determined by SDS-polyacrylamide slab gel electrophoresis was estimated to be 25 kDa. The purified enzyme showed maximum activity at pH 5 and at temperature of 55?C. Mg2+, Zn2+ and EDTA had an inhibitory effect on the activity of the acid phosphatase. Black gram seedling acid phosphatase was activated by K+, Cu2+ and Ba2+. The Km value of the enzyme was found to be 0.49 mM for pNPP as substrate.

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 Salivary apyrase of Rhodnius prolixus. Kinetics and purification

1986 ◽  
Vol 233 (3) ◽  
pp. 885-891 ◽  
Author(s):  
J J F Sarkis ◽  
J A Guimarães ◽  
J M C Ribeiro
Keyword(s):  
Atp Hydrolysis ◽  
Competitive Inhibition ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Rhodnius Prolixus ◽  
Exchange Chromatography ◽  
Phosphate Esters ◽  
Inorganic Pyrophosphatase ◽  
Blood Sucking ◽  
Specific Activities

The salivary apyrase activity of the blood-sucking bug Rhodnius prolixus was found to reside in a true apyrase (ATP diphosphohydrolase, EC 3.6.1.5) enzyme. The crude saliva was devoid of 5′-nucleotidase, inorganic pyrophosphatase, phosphatase and adenylate kinase activities. ATP hydrolysis proceeded directly to AMP and Pi without significant accumulation of ADP. Km values for ATP and ADP hydrolysis were 229 and 291 microM respectively. Ki values for ATP and ADP inhibition of ADP and ATP hydrolysis were not different from the Km values, and these experiments indicated competitive inhibition. Activities were purified 126-fold by combined gel filtration and ion-exchange chromatography procedures with a yield of 63%. The purified enzyme displayed specific activities of 580 and 335 mumol of Pi released/min per mg of protein for ATP and ADP hydrolysis respectively. The action of the purified enzyme on several phosphate esters indicates that Rhodnius apyrase is a non-specific nucleosidetriphosphate diphosphohydrolase.

scholarly journals Purification and Partial Characterization of β-Glucosidase from Fresh Leaves of Tea Plants (Camellia sinensis (L.) O. Kuntze)

2005 ◽  
Vol 37 (6) ◽  
pp. 363-370 ◽  
Author(s):  
Ye-Yun Li ◽  
Chang-Jun Jiang ◽  
Xiao-Chun Wan ◽  
Zheng-Zhu Zhang ◽  
Da-Xiang Li
Keyword(s):  
Specific Activity ◽  
Gel Filtration ◽  
Maximum Activity ◽  
Exchange Chromatography ◽  
Optimum Activity ◽  
Development Of Resistance ◽  
Sds Page ◽  
C 50 ◽  
Tea Plants

Abstractβ-Glucosidases are important in the formation of floral tea aroma and the development of resistance to pathogens and herbivores in tea plants. A novel β-glucosidase was purified 117-fold to homogeneity, with a yield of 1.26%, from tea leaves by chilled acetone and ammonium sulfate precipitation, ion exchange chromatography (CM-Sephadex C-50) and fast protein liquid chromatography (FPLC; Superdex 75, Resource S). The enzyme was a monomeric protein with specific activity of 2.57 U/mg. The molecular mass of the enzyme was estimated to be about 41 kDa and 34 kDa by SDS-PAGE and FPLC gel filtration on Superdex 200, respectively. The enzyme showed optimum activity at 50 °C and was stable at temperatures lower than 40 °C. It was active between pH 4.0 and pH 7.0, with an optimum activity at pH 5.5, and was fairly stable from pH 4.5 to pH 8.0. The enzyme showed maximum activity towards pNPG, low activity towards pNP-Galacto, and no activity towards pNP-Xylo.

scholarly journals Cathepsin B, the lysosomal thiol proteinase of calf liver

1969 ◽  
Vol 114 (4) ◽  
pp. 673-678 ◽  
Author(s):  
O. Snellman
Keyword(s):  
Cathepsin B ◽  
Ph Dependence ◽  
Gel Filtration ◽  
Chelating Agent ◽  
Thiol Group ◽  
Maximum Activity ◽  
Active Centre ◽  
Peptide Bonds ◽  
Hydrolysis Of ◽  
Thiol Proteinase

Cathepsin B from calf liver was obtained by a method involving preparation of a lysosomal–mitochondrial pellet and treatment of this pellet with acetone. The material was extracted with an acid buffer, pH4·0, and then precipitated from the extract with acetone. The precipitate was dissolved in phosphate buffer, pH7·4, and subjected to gel filtration on Sephadex G-200 and G-100. The cathepsin B emerged in a range of molecular weight much lower than 50000 as a well-defined component. The purity of this material was checked by electrophoresis. To obtain maximum activity the enzyme had to be activated with a chelating agent and a reducing agent (i.e. EDTA and cysteine). A number of different substrates were used. The enzyme was active for the hydrolysis of both peptide bonds and ester bonds and had approximately equal reactivity in the two cases. The pH-dependence of the hydrolysis was the same with both substrates. The binding of the substrates was half-maximal at pH4·5 and at pH6·8. A thiol group occurred in the active centre but this group ought to have a much higher pK than that found in this enzyme.

scholarly journals l(+)-Mandelate dehydrogenase from Rhodotorula graminis: purification, partial characterization and identification as a flavocytochrome b

1993 ◽  
Vol 293 (2) ◽  
pp. 455-460 ◽  
Author(s):  
M Yasin ◽  
C A Fewson
Keyword(s):  
Feedback Inhibition ◽  
Gel Filtration ◽  
Hydrophobic Interaction Chromatography ◽  
Maximum Activity ◽  
British Library ◽  
Terminal Sequence ◽  
Sds Page ◽  
Competitive Inhibitors ◽  
Lactate Dehydrogenases ◽  
Flavocytochrome B

L(+)-Mandelate dehydrogenase was purified to homogeneity from the yeast Rhodotorula graminis KGX 39 by a combination of (NH4)2SO4 fractionation, ion-exchange and hydrophobic-interaction chromatography and gel filtration. The amino-acid composition and the N-terminal sequence of the enzyme were determined. Comprehensive details of the sequence determinations have been deposited as Supplementary Publication SUP 50172 (4 pages) at the British Library Document Supply Centre, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1993) 289, 9. The enzyme is a tetramer as judged by comparison of its subunit M(r) value of 59,100 and native M(r) of 239,900, estimated by SDS/PAGE and gel filtration respectively. There is one molecule of haem and approx. one molecule of non-covalently bound FMN per subunit. 2,6-Dichloroindophenol, cytochrome c and ferricyanide can all serve as electron acceptors. L(+)-Mandelate dehydrogenase is stereospecific for its substrate. D(-)-Mandelate and L(+)-hexahydromandelate are competitive inhibitors. The enzyme has maximum activity at pH 7.9 and it has a pI value of 4.4. HgCl2 and 4-chloromercuribenzoate are potent inhibitors, but there is no evidence that the enzyme is subject to feedback inhibition by potential metabolic effectors. The evidence suggests that L(+)-mandelate dehydrogenase from R. graminis is a flavocytochrome b which is very similar to, and probably (at least so far as the haem domain is concerned) homologous with, certain well-characterized yeast L(+)-lactate dehydrogenases, and that the chief difference between them is their mutually exclusive substrate specificities.

scholarly journals Neutral maltase/glucoamylase from rabbit renal cortex

1989 ◽  
Vol 261 (1) ◽  
pp. 43-47 ◽  
Author(s):  
B Pereira ◽  
S Sivakami
Keyword(s):  
Brush Border ◽  
Brush Border Membrane ◽  
Renal Cortex ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Rabbit Kidney ◽  
Purification Procedure ◽  
Glucoamylase Activity ◽  
Maltase Activity ◽  
Triton X

Maltase activity (EC 3.2.1.20) was solubilized from rabbit kidney brush-border membrane by using 1.0% Triton X-100 and purified 230-fold with an overall recovery of 30%. The purification procedure makes use of heat precipitation, chromatography on DE-52 DEAE-cellulose and gel filtration on Sephacryl S-300. Rabbit kidney brush border exhibited glucoamylase activity with a maltase/glucoamylase ratio of 1.5:1 to 2.0:1. During purification the maltase and glucoamylase activities behaved identically. The Mr of the complex is 590,000, and it appears to be composed of eight identical subunits linked by disulphide bridges.

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