Characterization of Laccases and Peroxidases from Wood-Rotting Fungi (Family Coprinaceae)

1998 ◽  
Vol 64 (5) ◽  
pp. 1601-1606 ◽  
Author(s):  
Marion Heinzkill ◽  
Lisbeth Bech ◽  
Torben Halkier ◽  
Palle Schneider ◽  
Timm Anke
Keyword(s):  
Sodium Azide ◽  
Polyacrylamide Gel Electrophoresis ◽  
Ligninolytic Enzymes ◽  
Coprinus Cinereus ◽  
Tetraacetic Acid ◽  
Alkaline Ph ◽  
Ph Values ◽  
Native Polyacrylamide Gel Electrophoresis ◽  
Diethyldithiocarbamic Acid

Panaeolus sphinctrinus, Panaeolus papilionaceus, and Coprinus friesii are described as producers of ligninolytic enzymes. P. papilionaceus andP. sphinctrinus both produced a laccase. In addition,P. sphinctrinus produced a manganese peroxidase. C. friesii secreted a laccase and two peroxidases similar to the peroxidase of Coprinus cinereus. The purified laccases and peroxidases were characterized by broad substrate specificities, significant enzyme activities at alkaline pH values, and remarkably high pH optima. The two peroxidases of C. friesii remained active at pH 7.0 and 60°C for up to 60 min of incubation. The peroxidases were inhibited by sodium azide and ethylene glycol-bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid (EGTA), whereas the laccases were inhibited by sodium azide andN,N-diethyldithiocarbamic acid. As determined by native polyacrylamide gel electrophoresis and isoelectric focusing, all three fungi produced laccase isoenzymes.

scholarly journals Purification and Characterization of the Recombinant Thermus sp. Strain T2 α-Galactosidase Expressed in Escherichia coli

2001 ◽  
Vol 67 (4) ◽  
pp. 1601-1606 ◽  
Author(s):  
Mitsunori Ishiguro ◽  
Satoshi Kaneko ◽  
Atsushi Kuno ◽  
Yoshinori Koyama ◽  
Shigeki Yoshida ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Active Form ◽  
Gel Filtration ◽  
Amino Acid Sequences ◽  
Side Chain ◽  
Catalytic Function ◽  
Native Polyacrylamide Gel Electrophoresis

ABSTRACT The nucleotide sequence of the Thermus sp. strain T2 DNA coding for a thermostable α-galactosidase was determined. The deduced amino acid sequence of the enzyme predicts a polypeptide of 474 amino acids (M r, 53,514). The observed homology between the deduced amino acid sequences of the enzyme and α-galactosidase from Thermus brockianus was over 70%.Thermus sp. strain T2 α-galactosidase was expressed in its active form in Escherichia coli and purified. Native polyacrylamide gel electrophoresis and gel filtration chromatography data suggest that the enzyme is octameric. The enzyme was most active at 75°C forp-nitrophenyl-α-d-galactopyranoside hydrolysis, and it retained 50% of its initial activity after 1 h of incubation at 70°C. The enzyme was extremely stable over a broad range of pH (pH 6 to 13) after treatment at 40°C for 1 h. The enzyme acted on the terminal α-galactosyl residue, not on the side chain residue, of the galactomanno-oligosaccharides as well as those of yeasts and Mortierella vinacea α-galactosidase I. The enzyme has only one Cys residue in the molecule.para-Chloromercuribenzoic acid completely inhibited the enzyme but did not affect the mutant enzyme which contained Ala instead of Cys, indicating that this Cys residue is not responsible for its catalytic function.

Characterization of cellulase-free xylanases from the newly isolated Bacillus sp. strain BP-23

1993 ◽  
Vol 39 (12) ◽  
pp. 1162-1166 ◽  
Author(s):  
A. Blanco ◽  
F. I. J. Pastor
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Xylanase Activity ◽  
Hydrolytic Activity ◽  
Bacillus Sp ◽  
Birchwood Xylan ◽  
Xylanase Production ◽  
Ph Values ◽  
Wide Range ◽  
Molecular Masses

A Bacillus strain with xylanase activity has been isolated. Maximum xylanase production was obtained when the strain was cultured in media supplemented with birchwood xylan or rice straw; production was repressed by glucose and xylose. The optimal temperature and pH for xylanase activity were 45–50 °C and 5.5–7.5, respectively. Crude xylanase was highly stable at a wide range of pH values, retaining 100% of the activity after 24 h of incubation at 37 °C in buffer at pH 10.0. Analysis by polyacrylamide gel electrophoresis and zymogram techniques showed four xylanase activity bands with apparent molecular masses of 32, 48, 61, and 66 kDa. The most active of them (molecular mass 32 kDa) apparently corresponded to a xylanase with an isoelectric point (pI) of 9.3 in isoelectrofocusing gels developed as zymograms. Four other bands with xylanase activity were detected at pIs of 7.7, 5.6, 5.0, and 4.5. Analysis for carboxymethylcellulase activity revealed that only the band of 48 kDa and the band with a pI of 7.7 showed hydrolytic activity against the cellulosic substrate.Key words: Bacillus sp., xylanase, isolation.

Characterization of a cytosolic aminopeptidase from Encephalitozoon intestinalis

Parasitology ◽  
2002 ◽  
Vol 124 (1) ◽  
pp. 1-7 ◽  
Author(s):  
J. J. MILLERSHIP ◽  
C. L. CHAPPELL ◽  
P. C. OKHUYSEN ◽  
K. F. SNOWDEN
Keyword(s):  
Enzyme Activity ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Maximal Activity ◽  
Subcellular Fractionation ◽  
Aminopeptidase Activity ◽  
Encephalitozoon Intestinalis ◽  
Native Polyacrylamide Gel Electrophoresis ◽  
Aminopeptidase Inhibitors

Aminopeptidase activity was detected in Encephalitozoon intestinalis using a fluorometric assay. The aminopeptidase was capable of hydrolysing different amino acids bound to 7-amino-4-trifluoromethyl coumarin, with maximal activity against the amino acid, leucine. Aminopeptidase activity was localized in E. intestinalis spores and in intracellular stages. Enzymatic activity was inhibited by the traditional aminopeptidase inhibitors, bestatin and its analogue, nitrobestatin. Inhibition with the chelating agents, EDTA and 1,10-phenanthroline, suggested that the enzyme activity belongs to the metalloaminopeptidase class. Subcellular fractionation demonstrated that maximal enzyme activity was localized in the cytosolic fraction. Direct fluorogenic substrate analysis by native polyacrylamide gel electrophoresis estimated a molecular weight of 70·8 kDa. Direct fluorogenic analysis by polyacrylamide ampholyte gel electrophoresis indicated an isoelectric point of 4·8. The enzyme was both heat (>37 °C) and cold (<0 °C) labile with an optimal activity at pH 7·2. This is the first report characterizing a cytosolic aminopeptidase in microsporidia.

scholarly journals Isolation and characterization of desmosome-associated tonofilaments from rat intestinal brush border.

1981 ◽  
Vol 90 (1) ◽  
pp. 116-127 ◽  
Author(s):  
W W Franke ◽  
S Winter ◽  
C Grund ◽  
E Schmid ◽  
D L Schiller ◽  
...  
Keyword(s):  
Small Intestine ◽  
Brush Border ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Two Dimensional Gel Electrophoresis ◽  
Ph Values ◽  
Isolation And Characterization ◽  
Isoelectric Ph ◽  
Triton X

Epithelial cells of the small intestine, like those of other internal organs, contain intermediate-sized filaments immunologically related to epidermal prekeratin which are especially concentrated in the cell apex. Brush-order fractions were isolated from rat small intestine, and apical tonofilaments attached to desmosomal plaques and terminal web residues were prepared therefrom by extraction in high salt (1.5 M KCl) buffer and Triton X-100. The structure of these filaments was indistinguishable from that of epidermal tonofilaments and, as with epidermal prekeratin, filaments could be reconstituted from solubilized, denatured intestinal tonofilament protein. On SDS polyacrylamide gel electrophoresis of proteins of the extracted desmosome-tonofilament fractions, a number of typical brush-border proteins were absent or reduced, and enrichment of three major polypeptides of Mr 55,000, 48,000, and 40,000 was noted. On two-dimensional gel electrophoresis, the three enriched major polypeptides usually appeared as pairs of isoelectric variants, and the two smaller components (Mr 48,000, and 40,000) were relatively acidic (isoelectric pH values of 5.40 and below), compared to the Mr 55,000 protein which focused at pH values higher than 6.4. The tonofilament proteins were shown to be immunologically related to epidermal prekeratin by immunoreplica and blotting techniques using antibodies to bovine epidermal prekeratins. Similar major polypeptides were found in desmosome-attached tonofilaments from small intestine of mouse and cow. However, comparisons with epidermal tissues of cow and rat showed that all major polypeptides of intestinal tonofilaments were different from the major prekeratin polypeptides of epidermal tonofilaments. The results present the first analysis of a defined fraction of tonofilaments from a nonepidermal cell. The data indicate that structurally identical tonofilaments can be formed, in different types of cells, by different polypeptides of the cytokeratin family of proteins and that tonofilaments of various epithelia display tissue-specific patterns of their protein subunits.

scholarly journals Graphene Oxide/Fe-Based Composite Pre-Polymerized Coagulants: Synthesis, Characterization, and Potential Application in Water Treatment

2020 ◽  
Vol 6 (3) ◽  
pp. 44
Author(s):  
Athanasia K. Tolkou ◽  
Anastasios I. Zouboulis
Keyword(s):  
Graphene Oxide ◽  
Surface Water ◽  
Ph Value ◽  
Synthesis And Characterization ◽  
Alkaline Ph ◽  
Sem Images ◽  
Operational Conditions ◽  
Ph Values ◽  
First Time

This study presents for the first time the synthesis and characterization of GO (graphene oxide), PFSiC (polyferric silicate chloride), and hybrid GO-PFSiC derivatives, aiming to enhance synergistically the performance of coagulation, when applied for the treatment of water. The structure and the morphology of composite GO-PFSiC coagulants were studied in detail by the application of FTIR, XRD, and SEM characterization techniques. Furthermore, the proposed coagulants were applied for the treatment of simulated turbid surface water. The effects of the reagent’s dosage, pH value, and experimental/operational conditions on the coagulation efficiency, applied mainly for the removal of turbidity, were examined. The results, obtained from the FTIR and XRD measurements, showed the presence of a bond between the PFSiC and the GO surface, indicating that the PFSiC particles are distributed uniformly on the surface of graphene, which was also confirmed by the SEM images. Especially, the composite compound GO-PFSiC1.5-15-0.5 presents the most uniform distribution of iron on the surface of graphene oxide and exhibits the optimum coagulation efficiency, while it significantly reduces the turbidity for doses above 3–5 mg/L, i.e., achieving the respective legislation limit as proposed by WHO. Specifically, at the alkaline pH values (>7.9), the removal of turbidity reaches 96%. Consequently, the results of this study render these materials as potential coagulant agents for further research and applications, aiming to also achieve the co-removal of other water components.

scholarly journals Characterization of Pigments from the Berries of Syzygium caryophyllatum: Novel Source of Anthocyanins

2019 ◽  
Vol 7 (2) ◽  
pp. 291-297
Author(s):  
Premalatha Shetty ◽  
Aveena Roche ◽  
Sasidhar B Somappa
Keyword(s):  
Fluorescent Light ◽  
Alkaline Ph ◽  
Ph Values ◽  
Bluish Green ◽  
Wet Weight ◽  
Acidic Conditions ◽  
Anthocyanin Pigments ◽  
Green Coloration ◽  
Red Coloration

Extract was prepared from the pulp and peels of the Syzygium caryophyllatum berries. At pH ≤2, the extract showed prominent red coloration and at pH values above 7, the extract attained bluish-green coloration. These typical characteristics indicate presence of anthocyanin pigments in the berries. The yield of the anthocyanins was around 1.9mg per gram wet weight and, around 43% of the anthocyanin pigments appear to be present in polymerized forms.   At pH 1.0, the pigments absorbed maximally at 515nm and at pH 9.0, the pigments were found to absorb to a maximum extent in the range of 380-400nm.  The colors at alkaline pH were unstable, while the red coloration under acidic conditions was relatively stable when exposed to fluorescent light and temperature of 50°C. Mass spectrum of the extract showed predominance of malvidin and petunidine derivatives in the extract. Syzygium caryophyllatum berries can serve as reservoirs of anthocyanin pigments. Int. J. Appl. Sci. Biotechnol. Vol 7(2): 291-297

Heterozygous Abnormal Fibrinogen Osaka III with the Replacement of γ Arginine-275 by Histidine Has an Apparently Higher Molecular Weight γ-Chain Variant

1992 ◽  
Vol 68 (05) ◽  
pp. 534-538 ◽  
Author(s):  
Nobuhiko Yoshida ◽  
Shingi Imaoka ◽  
Hajime Hirata ◽  
Michio Matsuda ◽  
Shinji Asakura
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulfate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Tetraacetic Acid ◽  
Fibrin Monomer ◽  
Sds Page ◽  
Thrombotic Tendency ◽  
Chain Variant

SummaryCongenitally abnormal fibrinogen Osaka III with the replacement of γ Arg-275 by His was found in a 38-year-old female with no bleeding or thrombotic tendency. Release of fibrinopeptide(s) by thrombin or reptilase was normal, but her thrombin or reptilase time in the absence of calcium was markedly prolonged and the polymerization of preformed fibrin monomer which was prepared by the treatment of fibrinogen with thrombin or reptilase was also markedly defective. Propositus' fibrinogen had normal crosslinking abilities of α- and γ-chains. Analysis of fibrinogen chains on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the system of Laemmli only revealed the presence of abnormal γ-chain with an apparently higher molecular weight, the presence of which was more clearly detected with SDS-PAGE of fibrin monomer obtained by thrombin treatment. Purified fragment D1 of fibrinogen Osaka III also seemed to contain an apparently higher molecular weight fragment D1 γ remnant on Laemmli gels, which was digested faster than the normal control by plasmin in the presence of [ethy-lenebis(oxyethylenenitrilo)]tetraacetic acid (EGTA).

Prothrombin Metz : Purification and Characterization of a Variant of Human Prothrombin

1979 ◽  
Author(s):  
M Ribieto ◽  
J Elion ◽  
D Labie ◽  
F Josso
Keyword(s):  
Molecular Weight ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Factor Xa ◽  
Polyacrylamide Gel ◽  
Cleavage Pattern ◽  
Purification And Characterization ◽  
Double Immunodiffusion ◽  
The Family

For the purification of the abnormal prothrombin (Pt Metz), advantage has been taken of the existence in the family of three siblings who, being double heterozygotes for Pt Metz and a hypoprothrombinemia, have no normal Pt. Purification procedures included barium citrate adsorption and chromatography on DEAE Sephadex as for normal Pt. As opposed to some other variants (Pt Barcelona and Madrid), Pt Metz elutes as a single symetrical peak. By SDS polyacrylamide gel electrophoresis, this material is homogeneous and appears to have the same molecular weight as normal Pt. Comigration of normal and abnormal Pt in the absence of SDS, shows a double band suggesting an abnormal charge for the variant. Pt Metz exhibits an identity reaction with the control by double immunodiffusion. Upon activation by factor Xa, Pt Metz can generate amydolytic activity on Bz-Phe-Val-Arg-pNa (S2160), but only a very low clotting activity. Clear abnormalities are observed in the cleavage pattern of Pt Metz when monitored by SDS gel electrophoresis. The main feature are the accumulation of prethrombin l (Pl) and the appearance of abnormal intermediates migrating faster than Pl.

PURIFICATION AND PARTIAL CHEMICAL CHARACTERIZATION OF SHEEP NEUROPHYSIN

1973 ◽  
Vol 74 (2) ◽  
pp. 226-236 ◽  
Author(s):  
Michel Chrétien ◽  
Claude Gilardeau
Keyword(s):  
Amino Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Chemical Characterization ◽  
Terminal Amino Acid ◽  
Amino Acid Determination ◽  
Pituitary Glands ◽  
Terminal Amino ◽  
Partial Chemical ◽  
Acid Determination

ABSTRACT A protein isolated from ovine pituitary glands has been purified, and its homogeneity assessed by NH2- and COOH-terminal amino acid determination, ultracentrifugation studies, and polyacrylamide gel electrophoresis after carboxymethylation. Its chemical and immunochemical properties are closely similar to those of beef and pork neurophysins, less similar to those of human neurophysins. It contains no tryptophan (like other neurophysins) or histidine (like all except bovine neurophysin-I and human neurophysins). It has alanine at the NH2-terminus and valine at the COOH-terminus. Its amino acid composition is similar to, but not identical with those of porcine and bovine neurophysins.

Sign in / Sign up

Export Citation Format

Share Document