Extracellular alkaline phosphatase from marine bacteria: purification and properties of extracellular phosphatase from a marine Pseudomonas sp.

1980 ◽  
Vol 26 (7) ◽  
pp. 833-838 ◽  
Author(s):  
Hiromi Kobori ◽  
Nobuo Taga
Keyword(s):  
Molecular Weight ◽  
Alkaline Phosphatase ◽  
Enzyme Activity ◽  
Marine Bacteria ◽  
Divalent Cations ◽  
Maximal Activity ◽  
Pseudomonas Sp ◽  
Purification And Properties ◽  
Extracellular Alkaline Phosphatase ◽  
Increased Pressure

Extracellular alkaline phosphatase produced by a marine Pseudomonas was purified to electrophoretic homogeneity. The molecular weight of the enzyme was estimated to be 100 000. The enzyme had maximal activity at pH 11.5. The enzyme was completely inhibited by 1 mM EDTA. However, divalent cations reversed the enzyme inhibition and their order of effectiveness on the reaction was Zn2+ > Ca2+ > Mn2+ > Mg2+ > Sr2+ > Co2+. The enzyme activity was affected by the species of anion whose order of effectiveness was demonstrated to follow the lyotrophic series, Cl− > Br− > NO3−> ClO4− > SCN−. The activity of phosphatase was accelerated linearly by increased pressure until up to 1000 atm (1 atm = 101.325 kPa), and the enzyme activity at 1000 atm was 3.2 times higher than that at 1 atm.

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+.

PURIFICATION AND PROPERTIES OF A PROTEASE FROM PENICILLIUM CYANEO-FULVUM

1960 ◽  
Vol 38 (1) ◽  
pp. 969-980 ◽  
Author(s):  
Kartar Singh ◽  
S. M. Martin
Keyword(s):  
Molecular Weight ◽  
Enzyme Activity ◽  
Proteolytic Enzyme ◽  
Ethylenediaminetetraacetic Acid ◽  
Alkaline Proteases ◽  
Plasma Albumin ◽  
Metal Chelating ◽  
Bovine Plasma ◽  
Purification And Properties ◽  
Optimum Ph

A proteolytic enzyme present in culture filtrates of Penicillium cyaneo-fulvum was purified approximately 100-fold. In the ultracentrifuge the enzyme behaved as a homogeneous protein, but on electrophoresis some contamination was apparent. The molecular weight of the enzyme was estimated to be about 45,000.The protease hydrolyzed casein and denatured haemoglobin, gelatin, and native bovine plasma albumin but not native or denatured ovalbumin. It also coagulated milk. The optimum pH for caseolysis was 9.5 to 11.0. Metal chelating- and sulphydryl-reagents did not affect enzyme activity but zinc and mercurous ions inhibited the enzyme, the inhibition being reversed with ethylenediaminetetraacetic acid (EDTA). Soybean trypsin inhibitor was without effect on the enzyme whereas ovomucoid inhibited the enzyme. Although it is similar in some respects to other alkaline proteases, the P. cyaneo-fulvum enzyme does not appear to be identical with any one of them.

Purification and some properties of uridine diphosphate N-acetylglucosamine pyrophosphorylase from Neurospora crassa

1979 ◽  
Vol 25 (12) ◽  
pp. 1381-1386 ◽  
Author(s):  
Kenji Yamamoto ◽  
Mitsuaki Moriguchi ◽  
Hiroyasu Kawai ◽  
Tatsurokuro Tochikura
Keyword(s):  
Molecular Weight ◽  
Enzyme Activity ◽  
Neurospora Crassa ◽  
Isoelectric Point ◽  
Enzyme Preparation ◽  
Divalent Cations ◽  
Gel Filtration ◽  
Maximum Activity ◽  
Uridine Diphosphate ◽  
Chromatographic Techniques

Uridine diphosphate N-acetylglucosamine pyrophosphorylase (EC. 2.7.7.23) of Neurospora crassa has been purified approximately 210-fold with dithiothreitol as the stabilizing agent by use of chromatographic techniques. The enzyme preparation appeared to be homogeneous when subjected to electrophoresis. The molecular weight was estimated as approximately 37 000 by gel filtration. The enzyme had an isoelectric point around pH 4.4.Maximum activity of the enzyme was observed at pH7.5. The enzyme required Mg2+, which may be replaced by other divalent cations such as Mn2+ and Co2+ for lesser degrees of effectiveness. The enzyme was strictly specific for UDP-N-acetylglucosamine as the substrate. The estimated values of Km were 2.2 mM for UDP-N-acetylglucosamine and 5.4 mM for inorganic pyrophosphate.The enzyme activity was highly stimulated by the addition of dithiothreitol or dithioerythritol but was lost by sulfhydryl inhibitory reagents.

scholarly journals Purification and properties of a ribosomal casein kinase from rabbit reticulocytes

1977 ◽  
Vol 165 (3) ◽  
pp. 511-518 ◽  
Author(s):  
O G Issinger
Keyword(s):  
Molecular Weight ◽  
Enzyme Activity ◽  
Gradient Analysis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Casein Kinase ◽  
Purification And Properties ◽  
Casein Kinases ◽  
Acidic Proteins ◽  
Rabbit Reticulocytes

A casein kinase was isolated and purifed from rabbit reticulocytes. About 90% of the enzyme activity co-sedimented with the ribosomal fraction, whereas about 10% of the enzyme activity was found in the ribosome-free supernatant. Both casein kinases (the ribosome-bound enzyme as well as the free enzyme) showed identical activity and the same molecular weight. On sodium dodecyl sulphate/polyacrylamide-gel electrophoresis a single band of about 70 000 mol.wt. was observed. Sucrose-gradient analysis, however, showed that the enzyme activity sedimented with a s20,w of approx. 7.5S. This observation suggested that the casein kinase is a dimer composed of subunits of identical molecular weight. The enzyme utilizes GTP as well as ATP as a phosphoryl donor. It preferentially phosphorylates acidic proteins, in particular the model substrates casein and phosvitin. Casein kinase is cyclic AMP-indepenoent. The Km values for ATP and GTP with phosvitin as a substrate were determined as 1.2 and 8.8 micrometer respectively.

Purification and properties of phosphatidylgkycerophosphate synthetase from mammalian liver mitochondria

1978 ◽  
Vol 56 (6) ◽  
pp. 414-419 ◽  
Author(s):  
W. C. McMurray ◽  
E. C. Jarvis
Keyword(s):  
Divalent Cations ◽  
Maximal Activity ◽  
Liver Mitochondria ◽  
Mitochondrial Membranes ◽  
Purification And Properties ◽  
Standard Assay ◽  
Mammalian Liver ◽  
Triton X ◽  
Cytidine Diphosphate ◽  
Additional Stimulation

The enzyme which catalyzes the synthesis of phosphatidylglycerophosphate from sn-glycerol-3-phosphate and cytidine diphosphate diacylglycerol was released from rat or pig liver mitochondrial membranes by extraction with Triton X-100 or Nonidet P-40. The detergent-extracted enzyme, like the activity of intact mitochondria, did not require added cations or lipids.The Triton extracts were fractionated by column chromatography on Bio-Gel A-1.5. The fractions obtained from the columns exhibited little activity in the standard assay system unless divalent cations were included. Additional stimulation (about twofold) was observed in the presence of added phospholipids.The cation requirement of the purified enzyme was relatively nonspecific with Mg2+, Ba2+, or Ca2+ providing maximal activity in the 10 mM range. Either Mn2+ or Co2+ were stimulatory at somewhat lower concentrations but higher concentrations were inhibitory. Other cations such as Cd2+, Zn2+, Hg2+, or Cu2+ were ineffective as cofactors, and in the presence of Mg2+ inhibited the reaction at concentrations greater than 0.5 mM.The phospholipid stimulation was obtained specifically with phosphatidylethanolamines from natural or synthetic sources. Other diacylglycerophosphatides or lysophosphatides including lysophosphatidylethanolamine were ineffective.

Purification and Characterization of a Bioscouring Pectate Lyase from Paenibacillus sp. WZ008 with High Activity on Pectin

2012 ◽  
Vol 441 ◽  
pp. 457-461
Author(s):  
Xiang Xian Ying ◽  
Li Na Chen ◽  
Mei Lan Yu ◽  
Qun Xue ◽  
Zhao Wang
Keyword(s):  
Molecular Weight ◽  
Enzyme Activity ◽  
High Activity ◽  
Culture Supernatant ◽  
Pectate Lyase ◽  
Maximal Activity ◽  
Polygalacturonic Acid ◽  
Purification And Characterization ◽  
Optimum Activity

An extracellular pectate lyase was purified from the culture supernatant of Paenibacillus sp. WZ008 grown in the pectin-containing medium. The enzyme was purified to homogeneity in three steps and found to have a molecular weight of around 45 kDa. Highly methylated pectin was the optimum substrate in the case of no Ca2+ addition while the enzyme exhibited the maximal activity on polygalacturonic acid in the presence of 4 mM Ca2+. The purified enzyme demonstrated the optimum activity at a temperature range of 55-60°C and pH 9.6. The Ca2+ ion enhanced the enzyme activity but Mn2+, Ba2+ and EDTA strongly inhibited it.

scholarly journals Studies on the properties of cow's-milk tributyrinases and their interaction with milk proteins

1966 ◽  
Vol 101 (3) ◽  
pp. 651-660 ◽  
Author(s):  
WK Downey ◽  
P Andrews
Keyword(s):  
Molecular Weight ◽  
Alkaline Phosphatase ◽  
Enzyme Activity ◽  
Sodium Chloride ◽  
Xanthine Oxidase ◽  
Pancreatic Lipase ◽  
Wheat Germ ◽  
Milk Proteins ◽  
Casein Micelles ◽  
Bivalent Cation

1. The tributyrinases in milk are mainly associated with casein micelles. Dilution or addition of sodium chloride increases the enzyme activity, probably by dissociating the micelle-tributyrinase complexes. 2. Tributyrinase activities of milks activated by dilution and sodium chloride addition were in the range 0.2-1.7muequiv. of acid liberated/ml. of milk/min. from tributyrin emulsion at pH8.5 and 25 degrees . The enzymes have a bivalent-cation requirement for full activity and are rather unstable when separated from casein. 3. Ultracentrifugation of skim milks containing sodium chloride (0.75m) gave preparations low in casein but containing about 70% of the milk tributyrinases. The tributyrinases in such preparations appear to be bound in complexes of molecular weight about 350000. Dilution may result in dissociation to give the free enzymes. 4. Pancreatic lipase also formed complexes with casein micelles, but wheat-germ esterase, xanthine oxidase, milk alkaline phosphatase and other enzymes did not.

Purification and properties of a nonspecific β-fructofuranosidase (inulinase) from the mushroom Panaeolus papillonaceus

1987 ◽  
Vol 33 (6) ◽  
pp. 520-524 ◽  
Author(s):  
Khana Mukherjee ◽  
S. Sengupta
Keyword(s):  
Molecular Weight ◽  
Enzyme Activity ◽  
Activity Ratio ◽  
Optimum Temperature ◽  
Ph Optimum ◽  
Purification And Properties ◽  
Inulinase Activity ◽  
Wide Ph Range ◽  
Total Molecular Weight ◽  
Ph Range

A nonspecific β-fructofuranosidase (inulinase) was purified to electrophoretic homogeneity from the culture filtrate of the mushroom Panaeolus papillonaceus. The enzyme is the first purified from a basidiomycete and consists of two subunits with a total molecular weight of 116 000. It is most active on sucrose, then on raffinose, stachyose, and inulin, in decreasing order. The sucrase/inulinase activity ratio (S/I) is 5.7. Fructose was detected as the liberated sugar from raffinose, stachyose, and inulin. The enzyme is highly thermostable with an optimum temperature range of 60–65 °C and a pH optimum of 6.0. The enzyme is stable over the pH range 4–10, and is also active over a wide pH range, exhibiting 50% activity even at pH 8.5. Iodoacetate, azide, and EDTA, at 20 mM concentration, and 1% (w/v) SDS have no effect on enzyme activity, whereas Ag+ and Hg2+ at 2 mM are highly inhibitory.

Molecular characterization of an endo-type chitosanase from the fish pathogenRenibacteriumsp. QD1

2014 ◽  
Vol 94 (4) ◽  
pp. 681-686 ◽  
Author(s):  
Peichuan Xing ◽  
Dan Liu ◽  
Wen-Gong Yu ◽  
Xinzhi Lu
Keyword(s):  
Molecular Weight ◽  
Enzyme Activity ◽  
Specific Activity ◽  
Fermentation Broth ◽  
Maximal Activity ◽  
Optimum Temperature ◽  
Sds Page ◽  
Ph Range ◽  
Tlc Analysis

Renibacteriumsp. QD1, a bacteria strain capable of hydrolysing chitosan, was isolated from the homogenate of small crabs. An extracellular chitosanase, Csn-A, was purified from the QD1 fermentation broth. The enzyme was purified to homogeneity, with a yield of eight-fold, 67% recovery and a specific activity of 1575 U/mg proteins. The molecular weight of Csn-A was estimated to be 26.1 kDa by SDS-PAGE. Unlike other chitosanases, the purified Csn-A displayed maximal activity at a pH range of 5.3–6.5, and it was stable in a broad pH range of 5.0–10.0. The optimum temperature for chitosanlytic activity was 55°C. The enzyme activity was strongly stimulated by Mn2+but inhibited by Fe3+, Cu2+, Al3+, Zn2+and SDS. TLC analysis demonstrated that Csn-A hydrolysed N-deacetylated polymeric glucosamines into chito-biose and -triose in an endo-type manner. The amino acid seuquence of Csn-A showed close identity with an uncharacterized chitosanase of strain ATCC33209.

PURIFICATION AND PROPERTIES OF A PROTEASE FROM PENICILLIUM CYANEO-FULVUM

1960 ◽  
Vol 38 (9) ◽  
pp. 969-980 ◽  
Author(s):  
Kartar Singh ◽  
S. M. Martin
Keyword(s):  
Molecular Weight ◽  
Enzyme Activity ◽  
Proteolytic Enzyme ◽  
Ethylenediaminetetraacetic Acid ◽  
Alkaline Proteases ◽  
Plasma Albumin ◽  
Metal Chelating ◽  
Bovine Plasma ◽  
Purification And Properties ◽  
Optimum Ph

A proteolytic enzyme present in culture filtrates of Penicillium cyaneo-fulvum was purified approximately 100-fold. In the ultracentrifuge the enzyme behaved as a homogeneous protein, but on electrophoresis some contamination was apparent. The molecular weight of the enzyme was estimated to be about 45,000.The protease hydrolyzed casein and denatured haemoglobin, gelatin, and native bovine plasma albumin but not native or denatured ovalbumin. It also coagulated milk. The optimum pH for caseolysis was 9.5 to 11.0. Metal chelating- and sulphydryl-reagents did not affect enzyme activity but zinc and mercurous ions inhibited the enzyme, the inhibition being reversed with ethylenediaminetetraacetic acid (EDTA). Soybean trypsin inhibitor was without effect on the enzyme whereas ovomucoid inhibited the enzyme. Although it is similar in some respects to other alkaline proteases, the P. cyaneo-fulvum enzyme does not appear to be identical with any one of them.

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