Extracellular neuraminidase of Streptomyces albus

1972 ◽  
Vol 18 (7) ◽  
pp. 1007-1014 ◽  
Author(s):  
Marjory K. Myhill ◽  
Thomas M. Cook
Keyword(s):  
Gel Filtration ◽  
Thiobarbituric Acid ◽  
Enzyme Concentration ◽  
Divalent Metal Ions ◽  
Ph Optimum ◽  
Reactive Material ◽  
Culture Filtrates ◽  
Streptomyces Albus ◽  
Excess Substrate ◽  
Fetal Bovine

Extracellular neuraminidase activity (mucopolysaccharide N-acetylneuraminylhydrolase, EC.3.2.1.18) was detected in culture filtrates of Streptomyces albus MA-390, S. albus NCTC 7807, and S. albidoflaviis, but not S. albus ATCC 3004, S. roseochromogenes ATCC 13400, or S. venezuelae ATCC 10595. Neuraminidase of S. albus MA-390, purified 430-fold from crude culture filtrates, cleaved N-acetylneuraminyllactose and released thiobarbituric acid-reactive material from various glycoproteins. With excess substrate the reaction rate was proportional to enzyme concentration. The enzyme was not stimulated by divalent metal ions and was not inhibited by ethylenediaminetetraacetate. Streptomyces albus MA-390 neuraminidase was heat labile and activity was lost rapidly at temperatures of 40°C and above. With N-acetylneuraminyllactose as substrate optimal activity occurred at pH 6.0, but with fetal bovine serum the pH optimum was 3.5. The molecular weight of the enzyme was estimated by gel filtration to be in the range of 40 000 to 45 000.

THE WHEAT LEAF PHOSPHATASES: V. SOME PROPERTIES OF THE ENZYME SYSTEM HYDROLYZING β-GLYCEROPHOSPHATE IN CRUDE JUICE PREPARATIONS

1963 ◽  
Vol 41 (1) ◽  
pp. 113-120 ◽  
Author(s):  
D. W. A. Roberts
Keyword(s):  
Enzyme System ◽  
Enzyme Concentration ◽  
Ph Optimum ◽  
Wheat Leaf ◽  
Excess Substrate

Wheat leaf press juice contains an enzyme that has β-glycerophosphatase activity, which has a pH optimum close to pH 5.7. The enzyme is inhibited by orthophosphate, pyrophosphate, and 10−4 M fluoride. Fluoride inhibition can be abolished by thorough dialysis. Fluoride partially protects the enzyme from denaturation by heat.Enzyme kinetics shows that the log of the enzyme concentration is proportional to the log of the rate of liberation of orthophosphate from the substrate in the presence of excess substrate (0.2 M to 0.6 M) at pH 5.7. This observation can be used for quantitation of the enzyme.

THE WHEAT LEAF PHOSPHATASES: V. SOME PROPERTIES OF THE ENZYME SYSTEM HYDROLYZING β-GLYCEROPHOSPHATE IN CRUDE JUICE PREPARATIONS

1963 ◽  
Vol 41 (1) ◽  
pp. 113-120 ◽  
Author(s):  
D. W. A. Roberts
Keyword(s):  
Enzyme System ◽  
Enzyme Concentration ◽  
Ph Optimum ◽  
Wheat Leaf ◽  
Excess Substrate

Wheat leaf press juice contains an enzyme that has β-glycerophosphatase activity, which has a pH optimum close to pH 5.7. The enzyme is inhibited by orthophosphate, pyrophosphate, and 10−4 M fluoride. Fluoride inhibition can be abolished by thorough dialysis. Fluoride partially protects the enzyme from denaturation by heat.Enzyme kinetics shows that the log of the enzyme concentration is proportional to the log of the rate of liberation of orthophosphate from the substrate in the presence of excess substrate (0.2 M to 0.6 M) at pH 5.7. This observation can be used for quantitation of the enzyme.

scholarly journals cis, cis-Muconate cyclase from Trichosporon cutaneum

1980 ◽  
Vol 191 (1) ◽  
pp. 37-43 ◽  
Author(s):  
A Gaal ◽  
H Y Neujahr
Keyword(s):  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Polyacrylamide Gel ◽  
Trichosporon Cutaneum ◽  
Thiol Groups ◽  
Divalent Metal Ions ◽  
Ph Optimum

The inducible enzyme catalysing the conversion of cis, cis-muconate to (+)-muconolactone was purified 300-fold from the yeast Trichosporon cutaneum, grown on phenol. The enzyme has a sharp pH optimum at pH 6.6. It reacts also with several monohalogen derivatives and with one monomethyl derivative of cis, cis-muconate, but not with cis, trans- or trans, trans-muconate or 3-carboxy-cis, cis-muconate. In contrast with the corresponding enzymes in bacteria, the yeast enzyme does not require added divalent metal ions for activity and is not inhibited by EDTA. The purified enzyme can be resolved into two peaks by isoelectric focusing. The two forms have pI 4.58 (cis, cis-muconate cyclase I) and pI 4.74 (cis, cis-muconate cyclase II), respectively. Each of these is homogenous on polyacrylamide-gel electrophoresis in the absence or presence of sodium dodecyl sulphate. The two enzyme forms have the same molecular weight (50000) as determined by gel filtration and by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. They have the same Km value (25 microM) for cis, cis-muconate. They differ with respect to their content of free thiol groups. cis, cis-Muconate cyclase I contains one thiol group, essential for activity, but relatively stable upon storage. cis, cis-Muconate cyclase II contains two thiol groups that are readily oxidized during storage with concomitant loss of activity.

Isolation and Characterization of Plasminogen Activator from Pig Leucocytes

1974 ◽  
Vol 31 (01) ◽  
pp. 072-085 ◽  
Author(s):  
M Kopitar ◽  
M Stegnar ◽  
B Accetto ◽  
D Lebez
Keyword(s):  
Molecular Weight ◽  
Plasminogen Activator ◽  
Gel Electrophoresis ◽  
Gel Filtration ◽  
Ph Optimum ◽  
Isolation And Characterization ◽  
Ph Range ◽  
Inhibition Studies ◽  
Final Purification

SummaryPlasminogen activator was isolated from disrupted pig leucocytes by the aid of DEAE chromatography, gel filtration on Sephadex G-100 and final purification on CM cellulose, or by preparative gel electrophoresis.Isolated plasminogen activator corresponds No. 3 band of the starting sample of leucocyte cells (that is composed from 10 gel electrophoretic bands).pH optimum was found to be in pH range 8.0–8.5 and the highest pH stability is between pH range 5.0–8.0.Inhibition studies of isolated plasminogen activator were performed with EACA, AMCHA, PAMBA and Trasylol, using Anson and Astrup method. By Astrup method 100% inhibition was found with EACA and Trasylol and 30% with AMCHA. PAMBA gave 60% inhibition already at concentration 10–3 M/ml. Molecular weight of plasminogen activator was determined by gel filtration on Sephadex G-100. The value obtained from 4 different samples was found to be 28000–30500.

scholarly journals Thermostable glucose isomerase from psychrotolerant Streptomyces species

1970 ◽  
Vol 1 ◽  
pp. 6-10 ◽  
Author(s):  
Bidur Dhungel ◽  
Manoj Subedi ◽  
Kiran Babu Tiwari ◽  
Upendra Thapa Shrestha ◽  
Subarna Pokhrel ◽  
...  
Keyword(s):  
Specific Activity ◽  
Fold Increase ◽  
Glucose Isomerase ◽  
Enzyme Concentration ◽  
Maximal Velocity ◽  
Ph Optimum ◽  
Streptomyces Spp ◽  
Optimum Ph ◽  
Optimum Reaction ◽  
Sepharose 4B

Glucose isomerase (EC 5.3.1.5) was extracted from Streptomyces spp., isolated from Mt. Everest soil sample, and purified by ammonium sulfate fractionation and Sepharose-4B chromatography. A 7.1 fold increase in specific activity of the purified enzyme over crude was observed. Using glucose as substrate, the Michaelis constant (KM<) and maximal velocity (Vmax) were found to be 0.45M and 0.18U/mg. respectively. The optimum substrate (glucose) concentration, optimum enzyme concentration, optimum pH, optimum temperature, and optimum reaction time were 0.6M, 62.14μg/100μl, 6.9, 70ºC, and 30 minutes, respectively. Optimum concentrations of Mg2+ and Co2+ were 5mM and 0.5mM, respectively. The enzyme was thermostable with half-life 30 minutes at 100ºC.DOI: 10.3126/ijls.v1i0.2300 Int J Life Sci 1 : 6-10

Identification, purification, and characterization of phosphotyrosine-specific protein phosphatases from cultured chicken embryo fibroblasts

1984 ◽  
Vol 4 (6) ◽  
pp. 1003-1012
Author(s):  
R L Nelson ◽  
P E Branton
Keyword(s):  
Chicken Embryo ◽  
Protein Phosphatases ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Soluble Form ◽  
Ph Optimum ◽  
Cell Extracts ◽  
Phosphoprotein Phosphatases ◽  
Embryo Fibroblasts ◽  
Chicken Embryo Fibroblasts

Tyrosine phosphorylation catalyzed by a unique class of protein kinases is an important process in both normal cell proliferation and oncogenic transformation. In this study, phosphoprotein phosphatases specific for the dephosphorylation of phosphotyrosine residues were partially purified from secondary chicken embryo fibroblasts, using 32P-labeled immunoglobulin G phosphorylated by pp60src as substrate. Crude cell extracts contained ca. 70% of the activity in the soluble form and ca. 30% associated with a crude membrane fraction. The soluble activity was purified by using DEAE-cellulose and carboxymethyl cellulose column chromatography and gel filtration, and at least three enzyme species of apparent Mr 55,000 (pTPI), 50,000 (pTPII), and 95,000 (pTPIII)--comprising ca. 20, 45, and 35%, respectively, of the total activity--were resolved. All three enzymes possessed somewhat similar properties. They had a pH optimum of about 7.4, they were inhibited by Zn2+, vanadate, ATP, and ADP, and they were unaffected by divalent metal cations, EDTA, and F- under standard assay conditions employing a physiological ionic strength. These properties suggest that they represent a class of enzymes distinct from well-known phosphoseryl-phosphothreonyl-protein phosphatases and that dephosphorylation of phosphotyrosine-containing proteins may be carried out by a unique family of phosphoprotein phosphatases. Transformation by Rous sarcoma virus resulted in a small increase in phosphotyrosyl-protein phosphatase activity.

scholarly journals A Green Enzymatic Extraction Optimization and Oxidative Stability of Krill Oil from Euphausia Superba

Marine Drugs ◽  
2020 ◽  
Vol 18 (2) ◽  
pp. 82 ◽  
Author(s):  
Li Zhou ◽  
Fu Yang ◽  
Minghao Zhang ◽  
Jikai Liu
Keyword(s):  
Oxidative Stability ◽  
Ph Value ◽  
Thiobarbituric Acid ◽  
Euphausia Superba ◽  
Enzyme Concentration ◽  
Extraction Yield ◽  
Krill Oil ◽  
Tbars Values ◽  
Turbidity Increase

Krill oil enriched with polyunsaturated fatty acids is in the form of phospholipid. However, its application as a dietary supplement is limited, because of its rapid deterioration. Thus, this study aims to investigate the oxidative stability of krill oil extracted from Euphausia superba. Under optimal conditions (enzyme concentration 0.16%, enzymolysis time 2.9 h, and enzymolysis temperature of 45 °C) designed by response surface methodology, the extraction yield of krill oil is 86.02%. Five assays, including peroxide value (POV), thiobarbituric acid-reactive substances (TBARS), pH value, and turbidity were used to determine the oxidative stability of krill oil nanoliposomes during storage. Carboxymethyl chitosan (CMCS) nanoliposomes showed a significant reduction in POV and TBARS values, a prevention of pH value decrease and turbidity increase. This study indicated that CMCS nanoliposome can effectively improve the oxidative stability of krill oil during storage. Furthermore, the release profile in vitro illustrated that the controlled release of krill oil carried out by CMCS nanoliposomes is feasible.

Carboxymethylhydroxymuconic semialdehyde dehydrogenase in the 4-hydroxyphenylacetate catabolic pathway of Pseudomonas putida

1986 ◽  
Vol 64 (12) ◽  
pp. 1288-1293 ◽  
Author(s):  
Josefa M. Alonso ◽  
Amando Garrido-Pertierra
Keyword(s):  
Pseudomonas Putida ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Competitive Inhibitor ◽  
Catabolic Pathway ◽  
Ph Optimum ◽  
Semialdehyde Dehydrogenase ◽  
Standard Assay ◽  
Assay Conditions

5-Carboxymethyl-2-hydroxymuconic semialdehyde (CHMSA) dehydrogenase in the 4-hydroxyphenylacetate meta-cleavage pathway was purified from Pseudomonas putida by gel filtration, anion-exchange, and affinity chromatographies. Sodium dodecyl sulfate – polyacrylamide gel electrophoresis analysis suggested an approximate tetrameric molecular weight of 200 000. The purified enzyme showed a pH optimum at 7.8. The temperature–activity relationship for the enzyme from 27 to 45 °C showed broken Arrhenius plots with an inflexion at 36–37 °C. Under standard assay conditions, the enzyme acted preferentially with NAD. It could also catalyze the reduction with NADP (which had a higher Km), at 18% of the rate observed for NAD. The following kinetic parameters were found: Km(NAD) = 20.0 ± 3.6 μM, Km(CHMSA) = 8.5 ± 1.8 μM, and Kd(enzyme–NAD complex) = 7.8 ± 2.0 μM. The product NADH acted as a competitive inhibitor against NAD.

Experimental nephritis due to type specific streptococci. VI. Further characterization of type 12 nephrotoxin

1969 ◽  
Vol 15 (6) ◽  
pp. 555-561 ◽  
Author(s):  
P. W. Fardy ◽  
B. H. Matheson ◽  
R. W. Reed
Keyword(s):  
Active Material ◽  
Gel Filtration ◽  
Acute Glomerulonephritis ◽  
Acid Hydrolysate ◽  
Culture Filtrates ◽  
Experimental Nephritis ◽  
Chemical Studies ◽  
Group A ◽  
High Voltage Electrophoresis

Additional studies were undertaken on the nature of a nephrotoxic agent found in the culture filtrates of certain group A streptococci. A commercially available dehydrated medium proved satisfactory for the production of the active material. Gel filtration was used to divide polypeptide extracts, prepared from the dialyzable portion of culture filtrates, into two major fractions. One of these, representing the higher molecular weight components, contained most of the nephrotoxic activity as evidenced by the development of hypertension and acute glomerulonephritis in rabbits injected with this fraction.Physical and chemical studies indicated that the active fraction consisted of at least four polypeptide components separable by high voltage electrophoresis on paper. Automatic amino acid analysis of an acid hydrolysate of this fraction revealed 17 different amino acids. Carbohydrate was not detected by anthrone and orcinol tests.No relationship was established between this streptococcal nephrotoxic agent and other streptococcal constituents which have been implicated in acute glomerulonephritis.

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