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1.
Lignin peroxidase was purified (72-fold) from Acinetobacter calcoaceticus NCIM 2890. The purified lignin peroxidase (55–65 kDa) showed dimeric nature. The maximum enzyme activity was observed at
pH 1.0, between a broad temperature range of 50 and 70°C, at H 2O 2 concentration (40 mM) and the substrate concentration ( n-propanol, 100 mM). Purified lignin peroxidase was able to oxidize a variety of substrates including Mn 2+, tryptophan, mimosine, l-Dopa, hydroquinone, xylidine, n-propanol, veratryl alcohol, and ten textile dyes of various groups indicating as a versatile peroxidase. Most of the dyes
decolorized up to 90%. Tryptophan stabilizes the lignin peroxidase activity during decolorization of dyes. 相似文献
2.
Lignin polymers in bamboo ( Phyllostachys pubescens) were decomposed into polyphenols at high temperatures and oxidized for the introduction of quinone groups from peroxidase extracted from bamboo shoots and catalysis of UV. According to the results of FT-IR spectra analysis, neutral proteases (NPs) can be immobilized on the oxidized lignin by covalent bonding formed by amine group and quinone group. The optimum condition for the immobilization of NPs on the bamboo bar was obtained at pH 7.0, 40 °C, and duration of 4 h; the amount of immobilized enzyme was up to 5 mg g ?1 bamboo bar. The optimal pH for both free NP (FNP) and INP was approximately 7.0, and the maximum activity of INP was determined at 60 °C, whereas FNP presented maximum activity at 50 °C. The Km values of INP and FNP were determined as 0.773 and 0.843 mg ml ?1, respectively; INP showed a lower Km value and Vmax, than FNP, which demonstrated that INP presented higher affinity to substrate. Compared to FNP, INP showed broader thermal and storage stability under the same trial condition. With respect to cost, INP presented considerable recycling efficiency for up to six consecutive cycles. 相似文献
3.
The enzyme manganese peroxidase (MnP) is produced by numerous white-rot fungi to overcome biomass recalcitrance caused by
lignin. MnP acts directly on lignin and increases access of the woody structure to synergistic wood-degrading enzymes such
as cellulases and xylanases. Recombinant MnP (rMnP) can be produced in the yeast Pichia pastoris αMnP1-1 in fed-batch fermentations. The effects of pH and temperature on recombinant manganese peroxidase (rMnP) production
by P. pastoris αMnP1-1 were investigated in shake flask and fed-batch fermentations. The optimum pH and temperature for a standardized fed-batch
fermentation process for rMnP production in P. pastoris αMnP1-1 were determined to be pH 6 and 30 °C, respectively. P. pastoris αMnP1-1 constitutively expresses the manganese peroxidase ( mnp1) complementary DNA from Phanerochaete chrysosporium, and the rMnP has similar kinetic characteristics and pH activity and stability ranges as the wild-type MnP (wtMnP). Cultivation
of P. chrysosporium mycelia in stationary flasks for production of heme peroxidases is commonly conducted at low pH (pH 4.2). However, shake
flask and fed-batch fermentation experiments with P. pastoris αMnP1-1 demonstrated that rMnP production is highest at pH 6, with rMnP concentrations in the medium declining rapidly at
pH less than 5.5, although cell growth rates were similar from pH 4–7. Investigations of the cause of low rMnP production
at low pH were consistent with the hypothesis that intracellular proteases are released from dead and lysed yeast cells during
the fermentation that are active against rMnP at pH less than 5.5. 相似文献
4.
Alkaline thiol protease named Prot 1 was isolated from a culture filtrate of Botrytis cinerea. The enzyme was purified by ammonium sulfate fractionation, gel filtration, and ion-exchange chromatography. Thus, the enzyme
was purified to homogeneity with specific activity of 30-fold higher than that of the crude broth. The purified alkaline protease
has an apparent molecular mass of 43 kDa under denaturing conditions as estimated by sodium dodecyl sulfate-polyacrylamide
gel electrophoresis. The native molecular mass (45 kDa), determined by gel filtration, indicated that the alkaline protease
has a monomeric form. The purified protease was biochemically characterized. The enzyme is active at alkaline pH and has a
suitable and high thermostability. The optimal pH and temperature for activity were 9.0–10.0 and 60°C, respectively. This
protease was stable between pH 5.0 and 12.0. The enzyme retained 85% of its activity by treatment at 50°C over 120 min; it
maintained 50% of activity after 60 min of heating at 60°C. Furthermore, the protease retained almost complete activity after
4 wk storage at 25°C. The activity was significantly affected by thiol protease inhibitors, suggesting that the enzyme belongs
to the alkaline thiol protease family. With the aim on industrial applications, we focused on studying the stability of the
protease in several conditions. Prot 1 activity was not affected by ionic strength and different detergent additives, and,
thus, the protease shows remarkable properties as a biodetergent catalyst. 相似文献
5.
A Cyclodextrin (CDs) producing bacteria was isolated from waste of starch factory in Thailand and identified as Bacillus circulans by biochemical characterization and Paenibacillus sp. by 16S rRNA. The Paenibacillus grew and produced cyclodextrin glycosyltransferase (CGTase) at temperature range 37–45 °C. The optimum culturing conditions for highest CD-forming activity were pH 10.0 and 40 °C for 72 h in Horikoshi broth containing 0.5% soluble starch. The CGTase was partially purified by starch absorption, with 64% recovery and purification fold of 27. The optimum temperatures for dextrinizing and CD-forming activity were 70 and 50–55 °C. At the optimum temperature, the optimum pH for dextrinizing activity was 6.0, while CD-forming activity was 7.0. When the enzyme was incubated for 1 h at different temperatures, CD-forming activity retained its full activity up to 70 °C while dextrinizing activity dropped to 60%. Cyclodextrin products analyzed by HPLC was α:β=1:1, temperature of reaction mixture can affect the yield of CDs. 相似文献
6.
Arginase isolated from beef liver was covalently attached to a polyacrylamide bead support bearing carboxylic groups activated
by a water-soluble carbodiimide. The most favorable carbodiimide was N-cyclohexyl-Nt’-(methyl-2- p-nitrophenyl-2-oxoethyl) aminopropyl carbodiimide methyl bromide, but for practical purposes, N-cyclohexyl-Nt’-morpholinoethyl carbodiimide methyl tosylate was used. The optimal conditions for the coupling procedure were
determined. The catalytic activity of the immobilized arginase was 290–340 U/g solid or 2.9–3.4 U/mL wet gel. The pH optimum
for the catalytic activity was pH 9.5, the apparent temperature maximum was at 60°C and K mapp was calculated to be 0.37M L-arginine. Immobilization markedly improved the conformational stability of arginase. At 60°C,
the pH for maximal stability was found to be 8.0. The immobilized arginase was used for the production of L-ornithine and
D-arginine. 相似文献
7.
The effect of a number of environmental parameters (pH, temperature, carbon and nitrogen ratio of nutrient) on the production
of extracellular peroxidase enzymes by Streptomyces avermitilis UAH30 was examined. Maximum specific peroxidase activity (0.12 U/mg of protein) was obtained after 72 hours of 1 incubation
at 45‡C in a minimal salt medium (pH 7.5) containing 0.6% (w/v) yeast extract and 0.6% (w/v) xylan corresponding to a C:N
ratio of 4 to 1. A study of the effect of incubation on peroxidase activity showed that the enzyme was stable and active for
at least one hour after incubation at 50‡C while at higher temperatures the stability and activity of the peroxidase was reduced
such that at 60‡C the peroxidase activity has a half life of 20 min while at 80‡C the half life was reduced to 5 min. The
activation energy for deactivation as a result of thermal denaturation of the enzyme was calculated to be 80 ±7 kJ/mol. The
optimum pH for the activity occurred between a pH range of 6.5–8.5 with pKa 1 and pKa 2 of 5.1 ±0.1 and 9.7 ±0.1, respectively. The K m and V max for the peroxidase activity were determined to be 1.45 mM and 0.31 unit per mg protein respectively using 2,4dicholorophenol
(2,4-DCP) as a substrate. Characterization of the peroxidase activity revealed activity against L,3–4 dihydroxyphenylalanine
and guaiacol, while no inhibition of peroxidase activity could be detected with the haem inhibitors such as potassium cyanide
and sodium azide, suggesting the lack of haem component in the tertiary structure. Aspects of using the crude peroxidase preparation
in the pulp and paper industry are discussed. 相似文献
8.
Candida rugosa lipase was entrapped in silica sol-gel particles prepared by hydrolysis of methyltrimethoxysilane and assayed by p-nitrophenyl palmitate hydrolysis, as a function of pH and temperature, giving pH optima of 7.8 (free enzyme) and 5.0–8.0
(immobilized enzyme). The optimum temperature for the immobilized enzyme (50–55°C) was 19°C higher than for the free enzyme.
Thermal, operational, and storage stability were determined with n-butanol and bytyric acid, giving at 45°C a half-life 2.7 times greater for the immobilized enzyme; storage time was 21 d
at room temperature. For ester synthesis, the optimum temperature was 47°C, and high esterification conversions were obtained
under repeated batch cycles (half-life of 138 h). 相似文献
9.
An extracellular lipase was purified from the fermentation broth of Bacillus coagulans ZJU318 by CM-Sepharose chromatography, followed by Sephacryl S-200 chromatography. The lipase was purified 14.7-fold with
18% recovery and a specific activity of 141.1 U/mg. The molecular weight of the homogeneous enzyme was (32 kDa), determined
by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis. The enzyme activity was maximum at pH 9.0 and was stable
over a pH range of 7.0–10.0, and the optimum temperature for the enzyme reaction was 45°C. Little activity loss (6.2%) was
observed after 1 h of incubation at 40°C. However, the stability of the lipase decreased sharply at 50 and 60°C. The enzyme
activity was strongly inhibited by Ag + and Cu 2+, whereas EDTA caused no inhibition. SDS, Brij 30, and Tween-80 inhibited lipase, whereas Triton X-100 did not significantly
inhibit lipase activity. 相似文献
10.
Partially purified α-Galactosidase from Penicillium griseoroseum was immobilized onto modified silica using glutaraldehyde linkages. The effective activity of immobilized enzyme was 33%.
Free and immobilized α-galactosidase showed optimal activity at 45 °C and pH values of 5 and 4, respectively. Immobilized
α-galactosidase was more stable at higher temperatures and pH values. Immobilized α-galactosidase from P. griseoroseum maintained 100% activity after 24 h of incubation at 40 °C, while free enzyme showed only 32% activity under the same incubation
conditions. Defatted soybean flour was treated with free and immobilized α-galactosidase in batch reactors. After 8 h of incubation,
stachyose was completely hydrolyzed in both treatments. After 8 h of incubation, 39% and 70% of raffinose was hydrolyzed with
free and immobilized α-galactosidase respectively. Immobilized α-galactosidase was reutilized eight times without any decrease
in its activity. 相似文献
11.
The marine yeast strain Kodamea ohmeri BG3 isolated from the gut of a marine fish ( Hexagrammes otakii) was found to secrete a large amount of phytase into the medium. The crude phytase produced by this marine yeast showed the
highest activity at pH 5.0 and 65 °C. The optimal medium for phytase production contained oat 10.0 g/l, ammonium sulfate 15.0 g/l,
glucose 30 g/l, and NaCl 20.0 g/l, while the optimal cultivation conditions for phytase production were pH 5.0, a temperature
of 28 °C, and a shaking speed of 170 rpm. Under the optimal conditions, over 557.9 mU/ml of phytase activity was produced
within 72 h of fermentation at the shake flask level. This is a very high level of phytase activity produced by yeasts. We
think that the medium and process for phytase production by the marine yeast strain were very simple, and such marine yeast
from the gut of natural marine fish may have a potential application in the maricultural industry and marine environmental
protection. The results demonstrate that phytate was actively degraded by the crude phytase within a short period. 相似文献
12.
Coal polymers, water soluble at pH 3.5, were prepared from North Dakota lignite and German subbituminous coal in 35–61% yield. Gel permeation chromatography showed a major component of relatively narrow molecular weight range >75,000. The material did not dialyze through a 12,000-14,000 MW cutoff membrane under several conditions. Minor amounts of smaller fragments were present, but monomeric components were not detected. Incubation of soluble polymer with lignin peroxidase ofPhanerochaete chrysosporium caused substantial disappearance of the high molecular weight polymer and formation of smaller amounts of both higher and lower molecular weight components, but not of monomeric compounds. Addition of veratryl alcohol enhanced depolymerization. Coal polymer competitively inhibited veratryl alcohol oxidation by lignin peroxidase. 相似文献
13.
A gene-encoding alkaline phosphatase (AP) from thermophilic Geobacillus thermodenitrificans T2, termed Gtd AP, was cloned and sequenced. The deduced Gtd AP protein comprises 424 amino acids and shares a low homology with other known AP (<35% identity), while it exhibits the
conservation of the active site and structure element of Escherichia coli AP. The Gtd AP protein, without a predicted signal peptide of 30 amino acids, was successfully overexpressed in E. coli and purified as a hexa-His-tagged fusion protein. The pH and temperature optima for purified enzyme are 9.0 and 65 °C, respectively.
The enzyme retained a high activity at 45–60 °C, while it could be quickly inactivated by a heat treatment at 80 °C for 15 min,
exhibiting a half-life of 8 min at 70 °C. The K
m and V
max for pNPP were determined to be 31.5 μM and 430 μM/min at optimal conditions. A divalent cation is essential, with a combination
of Mg 2+ and Co 2+ or Zn 2+ preferred. The enzyme was strongly inhibited by 10 mM ethylenediaminetetraacetic acid (EDTA) and vanadate but highly resistant
to urea and dithiothreitol. The properties of Gtd AP make it suitable for application in molecular cloning or amplification. 相似文献
14.
This study investigated the optimization of the enzymatic processing conditions for polylactic acid (PLA) fibers using enzymes
consisting of lipases originating from different sources. The hydrolytic activity was evaluated taking into consideration
the pH, temperature, enzyme concentration, and treatment time. The structural change of the PLA fibers was measured in the
optimal treatment conditions. PLA fiber hydrolysis by lipases was maximized for lipase from Aspergillus niger at 40 °C for 60 min at pH 7.5 with 60% (owf) concentration, for lipase from Candida cylindracea at 40 °C for 120 min at pH 8.0 with 70% (owf) concentration, and for lipase from Candida rugosa at 45 °C for 120 min at pH 8.0 with 70% (owf) concentration. There was a change in protein absorbance of the treatment solution
before and after all lipase treatments. The analyses of the chemical structure change and structural properties of the PLA
due to lipase treatment was confirmed by tensile strength, differential scanning calorimetry, wide-angle X-ray scattering
diffractometry, Fourier transform infrared spectroscopy, and scanning electron microscopy. 相似文献
15.
Lignocellulosic wastes such as neem hull, wheat bran, and sugarcane bagasse, available in abundance, are excellent substrates
for the production of ligninolytic enzymes under solid-state fermentation by white-rot fungi. A ligninolytic enzyme system
with high activity showing enhanced decomposition was obtained by cocultivation of Pleurotus ostreatus and Phanerochaete chrysosporium on combinations of lignocellulosic waste. Among the various substrate combinations examined, neem hull and wheat bran wastes
gave the highest ligninolytic activity. A maximum production of laccase of 772 U/g and manganese peroxidase of 982 U/g was
obtained on d 20 and lignin peroxidase of 656 U/g on d 25 at 28±1 °C under solid-state fermentation. All three enzymes thus
obtained were partially purified by acetone fractionation and were exploited for decolorizing different types of acid and
reactive dyes. 相似文献
16.
The possibility of producing the biologically active material of the skin, ceramide, was studied using yeasts. The yeast strain
that produced the most ceramide, Saccharomyces cerevisiae (KCCM 50515), was selected, and the optimal conditions for ceramide production were determined using shakeflask culture and
batch fermentation. By measuring the production rate of ceramide at various pH values and temperatures, the optimal conditions
for ceramide production were found to be pH 6.0 and 30°C. When heat shock was applied to the cells for 1 h by increasing the
culture temperature from 30 to 40°C after cell growth, the amount of ceramide produced was increased 5.9-fold. A cell growth
and ceramide production model was developed with Monod kinetics and the Leudecking-Piret model. It showed that ceramide production
was increased when the cells were in the stationary phase. 相似文献
17.
The enzyme cellobiase Novozym 188, which is used for improving hydrolysis of bagasse with cellulase, was characterized in
its commercial available form and integrated kinetic models were applied to the hydrolysis of cellobiose. The specific activity
of this enzyme was determined for pH values from 3.0–7.0, and temperatures from 40–75°C, with cellobiose at 2 g/L. Thermal
stability was measured at pH 4.8 and temperatures from 40–70°C. Substrate inhibition was studied at the same pH, 50°C, and
cellobiose concentrations from 0.4–20 g/L. Product inhibition was determined at 50°C, pH 4.8, cellobiose concentrations of
2 and 20 g/L, and initial glucose concentration nearly zero or 1.8 g/L. The enzyme has shown the greatest specific activity,
17.8 U/mg, at pH 4.5 and 65°C. Thermal activation of the enzyme followed Arrhenius equation with the Energy of Activation
being equal to 11 kcal/mol for pH values 4 and 5. Thermal deactivation was adequately modeled by the exponential decay model
with Energy of Deactivation giving 81.6 kcal/mol. Kinetics parameters for substrate uncompetitive inhibition were: Km=2.42 m M, V
max=16.31 U/mg, Ks=54.2 m M. Substrate inhibition was clearly observed above 10 m M cellobiose. Product inhibition at the concentration studied has usually doubled the time necessary to reach the same conversion
at the lower temperature tested. 相似文献
18.
The enantiomeric separation of several basic drugs was investigated using copper(II)–clindamycin as a new chiral selector. The results show that the chiral selector allows high-resolution separation of some racemic basic drugs, including tropicamide, propranolol, sotalol, bisoprolol, epinephrine, esmolol, atenolol, and metoprolol. The enantioselectivity was influenced by parameters such as the type of metal ion, ratio of clindamycin and Cu(II), pH of the background electrolyte, clindamycin concentration, applied voltage, and capillary temperature. The optimal separation conditions were determined to be 20 mM clindamycin/10 mM Cu2+, pH 9.06, at 20 kV and 22 °C within 25 min. 相似文献
19.
A simple, accurate and reproducible capillary electrophoresis method with UV detection has been developed for the simultaneous determination of four iridoid glycosides, 6-O-methyl-catalpol, aucubin, harpagide, and harpagoside, in Scrophularia ningpoensis (Xuan-shen). The running buffer was 100 mM borate (pH 9.3) containing 20% methanol. Applied voltage was 20 kV and temperature was 25 °C. Diphylloside A was used as an internal standard (IS) and detection was at 200 nm. The effects on separation of buffer pH, buffer concentration, and organic modifiers were investigated. The extracts of S. ningpoensis were well separated within 45 min. 相似文献
20.
Chloroperoxidase from M usa paradisiaca stem juice has been purified to homogeneity using a concentration obtained by ultrafiltration and anion exchange chromatography on diethylaminoethyl (DEAE) cellulose. The purified enzyme gave a single protein band in SDS‐PAGE analysis corresponding to molecular mass of 43 kDa. The native PAGE analysis result has also given a single protein band, confirming the purity of the enzyme. The purified enzyme was chlorinated and brominated with monochlorodimedone, the substrate used for measuring the halogenating activity of chloroperoxidases. The Km and kcat values using monochlorodimedone as the substrate were 20 μM and 1.64 s ?1, respectively, giving a kcat/ Km value of 8.2 × 10 4 M ?1 s ?1. The pH and temperature optima of the chlorinating activity were 3.0 and 25°C, respectively. The Km values for the peroxidase activity using pyragallol and H 2O 2 as the variable substrates were 89 and 120 μM, respectively. The pH and temperature optima of the peroxidase activity using pyrogalllol as the substrate were the same as the pH and temperature optima of the halogenating activity. The peroxidase activity of the enzyme is competitively inhibited by sodium azide, indicating that it is a hemeperoxidase different from nonheme peroxidases. © 2012 Wiley Periodicals, Inc. Int J Chem Kinet 45: 92–100, 2013 相似文献
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