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1.
The present work analyzes the activity in decomposition of H 2O 2 using magnetite-immobilized catalase. The support of catalase is a glutaraldehyde-treated magnetite (Fe 3O 4). The data obtained in the H 2O 2 decomposition are analyzed. The fitting of the initial rate of the H 2O 2 decomposition versus hydrogen peroxide concentration data is discussed using a specific program for enzyme kinetics modeling (Leonora). The free catalase from Aspergillus niger (3.5 or 10 U/mL) does not show substrate inactivation up to 0.4 M H 2O 2. The immobilized catalase at low catalyst concentration shows substrate inhibition. Using 1 mg/mL of supported catalase the predicted maximum activity is higher than in the case of the free catalase at similar catalase concentration, although the optimum temperature is lower (40 °C versus 60 °C). 相似文献
2.
Bovine liver catalase was encapsulated in an aqueous phase of the phospholipid vesicle (liposome) to improve the stability of its tetrameric structure and activity. The catalase-containing liposomes (CALs) prepared were 30, 50 and 100 nm in mean diameters (CAL 30, CAL 50 and CAL 100, respectively). The CAL 100 included the types I, II and III based on the amounts of catalase encapsulated. The CAL 30, CAL 50 and CAL 100-I contained one catalase molecule per liposome, and the CAL 100-II and CAL 100-III on average 5.2 and 17 molecules, respectively. The storage stability of catalase in either CAL system was significantly increased compared to that of free catalase at 4 °C in a buffer of pH 7.4. At 55 °C, free catalase was much more deactivated especially with decreasing its concentration predominantly due to enhanced dissociation of catalase into subunits while it was so done at excessively high enzyme concentration mainly due to enhanced formation of catalase intermolecular aggregates. Among the three types of CAL 100, the CAL 100-II showed the highest thermal stability, indicating that an excess amount of catalase in the CAL 100-III was also disadvantageous to maintain an active form of the catalase even in liposome. In the CAL 100-III, however, the stability of catalase was significantly improved compared to that of free catalase at the same concentration. The CAL thermal stability was little affected by the liposome size as observed in the CAL 30, CAL 50 and CAL 100-I. An intrinsic tryptophan fluorescence of the catalase recovered from the CAL 100-II thermally treated at 55 °C revealed that a partially denatured catalase molecule was stabilized through its hydrophobic interaction with liposome membrane. This interaction depressed not only dissociation of catalase into subunits but also formation of an inactive intermolecular aggregate between the catalase molecules in a liposome. Furthermore, either type of CAL 100 showed a higher stability than free catalase in the successive decompositions of 10 mM H 2O 2 at 25 °C mainly because the H 2O 2 concentration was kept low inside liposomes due to the permeation barrier of the lipid membrane to H 2O 2. 相似文献
3.
The turning point between apoptosis and necrosis induced by hydrogen peroxide (H 2O 2) have been investigated using human T-lymphoma Jurkat cells. Cells treated with 50 μM H 2O 2 exhibited caspase-9 and caspase-3 activation, finally leading to apoptotic cell death. Treatment with 500 μM H 2O 2 did not exhibit caspase activation and changed the mode of death to necrosis. On the other hand, the release of cytochrome c from the mitochondria was observed under both conditions. Treatment with 500 μM H 2O 2, but not with 50 μM H 2O 2, caused a marked decrease in the intracellular ATP level; this is essential for apoptosome formation. H 2O 2-reducing enzymes such as cellular glutathione peroxidase (cGPx) and catalase, which are important for the activation of caspases, were active under the 500 μM H 2O 2 condition. Prevention of intracellular ATP loss, which did not influence cytochrome c release, significantly activated caspases, changing the mode of cell death from necrosis to apoptosis. These results suggest that ATP-dependent apoptosome formation determines whether H 2O 2-induced cell death is due to apoptosis or necrosis. 相似文献
4.
The effect of vitamin C (ascorbate) on oxidative DNA damage was examined by first incubating cells with dehydroascorbate, which boosts the intracellular concentration of ascorbate, and then exposing cells to H 2O 2. Oxidative DNA damage was estimated by the analysis of 5-hydroxy-2′-deoxycytidine (oh 5dCyd) and 8-oxo-7,8-dihydro-2′-deoxyguanosine (oxo 8dGuo). The presence of a high concentration of ascorbate (30 mM), compared to the absence of ascorbate in cells, when exposed to H 2O 2 (200 μM), resulted in a remarkable sensitization of oh 5dCyd from 2.7 ± 0.6 to 40.8 ± 6.1 lesions /10 6 dCyd (15-fold). In contrast, the level of oxo 8dGuo increased from 8.4 ± 0.4 to 12.1 ± 0.5 lesions/10 6 dGuo (50%). The formation of oh 5dCyd was also observed at lower concentrations of intracellular ascorbate and exogenous H 2O 2. Additional studies showed that replacement of H 2O 2 with tert-butyl hydroperoxide completely abolished damage, and that preincubation with iron and desferroxamine increased and decreased this damage, respectively. The latter studies suggest that a Fenton reaction is involved in the mechanism of damage. In conclusion, we report a novel model system in which ascorbate sensitizes H 2O 2-induced oxidative DNA damage in cells, leading to elevated levels of oh 5dCyd and oxo 8dGuo, with a strong bias toward the formation of oh 5dCyd. 相似文献
5.
The hydrothermal reactions of (Ph 4P)[VO 2Cl 2] and H 2C 2O 4 at 150 and 125°C yield (Ph 4P) 2[V 2O 2(H 2O) 2(C 2O 4) 3]·4H 2O (1) and (Ph 4P)[VOCl(C 2O 4)] (2), respectively. The structure of the molecular anion of 1 consists of a binuclear unit of oxovanadium(IV) octahedra bridged by a bisbidentate oxalate group. The VO 6 coordination geometry at each vanadium site is defined by a terminal oxo group, an aquo ligand, and four oxygen donors — two from the bisbidentate bridging oxalate and two from the terminal bidentate oxalate. The structure of 2 consists of discrete Ph 4P + cations occupying regions between [VOCl(C 2O 4)] −∞ spiral chains. The structure of the one-dimensional anionic chain exhibits V(IV) octahedra bridged by bisbidentate oxalate groups. Crystal data: 1·4H 2O, monoclinic P2 1/ n, A = 12.694(3), B = 12.531(3), C = 17.17(3) Å, β = 106.32(2)°, V = 2621.3(13) Å 3, Z = 2, Dcalc = 1.501 g cm −3, structure solution and refinement converged at a conventional residual of 0.0518; 2, tetragonal P4 3, A = 12.145(2), C = 15.991(3) Å, V = 2358.7(12) Å 3, Z = 4, R = 0.0452. 相似文献
6.
Yeast cytochrome c peroxidase (CCP) efficiently catalyzes the reduction of H 2O 2 to H 2O by ferrocytochrome c in vitro. The physiological function of CCP, a heme peroxidase that is targeted to the mitochondrial intermembrane space of Saccharomyces cerevisiae, is not known. CCP1-null-mutant cells in the W303-1B genetic background ( ccp1Δ) grew as well as wild-type cells with glucose, ethanol, glycerol or lactate as carbon sources but with a shorter initial doubling time. Monitoring growth over 10 days demonstrated that CCP1 does not enhance mitochondrial function in unstressed cells. No role for CCP1 was apparent in cells exposed to heat stress under aerobic or anaerobic conditions. However, the detoxification function of CCP protected respiring mitochondria when cells were challenged with H 2O 2. Transformation of ccp1Δ with ccp1W191F, which encodes the CCP W191F mutant enzyme lacking CCP activity, significantly increased the sensitivity to H 2O 2 of exponential-phase fermenting cells. In contrast, stationary-phase (7-day) ccp1Δ- ccp1W191F exhibited wild-type tolerance to H 2O 2, which exceeded that of ccp1Δ. Challenge with H 2O 2 caused increased CCP, superoxide dismutase and catalase antioxidant enzyme activities (but not glutathione reductase activity) in exponentially growing cells and decreased antioxidant activities in stationary-phase cells. Although unstressed stationary-phase ccp1Δ exhibited the highest catalase and glutathione reductase activities, a greater loss of these antioxidant activities was observed on H 2O 2 exposure in ccp1Δ than in ccp1Δ- ccp1W191F and wild-type cells. The phenotypic differences reported here between the ccp1Δ and ccp1Δ- ccp1W191F strains lacking CCP activity provide strong evidence that CCP has separate antioxidant and signaling functions in yeast. 相似文献
7.
The effect of Hb-I* phenotype on white muscle lactate dehydrogenease (LDH, E. C. 1.1.1.27) activity and buffering capacity was studied in Atlantic cod ( Gadus morhua), acclimated and measured at temperatures near their behavioral temperature preference. It was hypothesized that these conditions would optimize biochemical processes but no difference was found in LDH activity between the Hb-I* phenotype after 56 d of acclimation to 6 and 14°C. However, LDH activity was both mass- and temperature-dependent; mean activity was 162.2±5.0 and 275.9±6.4 IU g -1 wet mass (mean±SEM) at 6 and 14°C respectively and larger fish had the highest rate of enzyme activity. White muscle buffer capacity was unaffected by Hb-I* phenotype but higher in cod held at 14°C. 相似文献
8.
1. Single reduced methyl viologen (MV .+) acts as an electron donor in a number of enzyme systems. The large changes in extinction coefficient upon oxidation (λ max 600 nm; MV .+, = 1.3 · 10 4 M −1 · cm −1; oxidised form of methyl viologen (MV 2+), = 0.0) make it ideally suited to kinetic studies of electron transfer reactions using stopped-flow and standard spectrophotometric techniques. 2. A convenient electrochemical preparation of large amounts of MV.+ has been developed. 3. A commercial stopped-flow apparatus was modified in order to obtain a high degree of anaerobicity. 4. The reaction of MV.+ with O2 produced H2O2 (k > 5 · 106 M−1 · s−1, pH 7.5, 25 °C). H2O2 subsequently reacted with excess MV.+ (k = 2.3 · 103 M−1 · s−1, pH 7.5, 25 °C) to produce water. The kinetics of this reaction were complex and have only been interpreted over a limited range of concentrations. 5. The results support the theory that the herbicidal action of methyl viologen (Paraquat, Gramoxone) is due to H2O2 (or radicals derived from H2O2) induced damage of plant cell membrane. 相似文献
9.
New mixed metal complexes SrCu 2(O 2CR) 3(bdmap) 3 (R = CF 3 (1a), CH 3 (1b)) and a new dinuclear bismuth complex Bi 2(O 2CCH 3) 4(bdmap) 2(H 2O) (2) have been synthesized. Their crystal structures have been determined by single-crystal X-ray diffraction analyses. Thermal decomposition behaviors of these complexes have been examined by TGA and X-ray powder diffraction analyses. While compound 1a decomposes to SrF 2 and CuO at about 380°C, compound 1b decomposes to the corresponding oxides above 800°C. Compound 2 decomposes cleanly to Bi 2O 3 at 330°C. The magnetism of 1a was examined by the measurement of susceptibility from 5–300 K. Theoretical fitting for the susceptibility data revealed that 1a is an antiferromagnetically coupled system with g = 2.012(7), −2 J = 34.0(8) cm −1. Crystal data for 1a: C 27H 51N 6O 9F 9Cu 2Sr/THF, monoclinic space group P2 1/ m, A = 10.708(6), B = 15.20(1), C = 15.404(7) Å, β = 107.94(4)°, V = 2386(2) Å 3, Z = 2; for 1b: C 27H 60N 6O 9Cu 2Sr/THF, orthorhombic space group Pbcn, A = 19.164(9), B = 26.829(8), C = 17.240(9) Å, V = 8864(5) Å 3, Z = 8; for 2: C 22H 48O 11N 4Bi 2, monoclinic space group P2 1/ c, A = 17.614(9), B = 10.741(3), C = 18.910(7) Å, β = 109.99(3)°, V = 3362(2) Å 3, Z = 4. 相似文献
10.
The ability of oxidative stress to induce apoptosis (programmed cell death), and the effect of Trolox, a water soluble vitamin E analog, on this induction were studied in vitro in mouse thymocytes. Cells were exposed to oxidative stress by treating them with 0.5–10 μM hydrogen peroxide (H 2O 2) for 10 min, in phosphate-buffered saline supplemented with 0.1 mM ferrous sulfate. Cells were resuspended in RPMI 1640 medium with 10% serum and incubated at 37°C under 5% CO 2 in air. Electron microscopic studies revealed morphological changes characteritic of apoptosis in H 2O 2-treated fragmented the DNA in a manner typical of apoptotic cells, producing a ladder pattern of 200 base pair increments upon agarose gel electrophoresis. The percentage of DNA fragmentation (determined fluorometrically) increased with increasing doses of H 2O 2 and postexposure incubation times. Pre- or posttreatment of cells with Trolox reduced H 2O 2-induced DNA fragmentation to control levels and below. The results indicate that oxidative stress induces apoptosis in thymocytes, and this induction can be prevented by Trolox, a powerful inhibitor of membrane damage. 相似文献
11.
Heme catalases are considered to degrade two molecules of H 2O 2 to two molecules of H 2O and one molecule of O 2 employing the catalatic cycle. We here studied the catalytic behaviour of bovine liver catalase at low fluxes of H 2O 2 (relative to catalase concentration), adjusted by H 2O 2-generating systems. At a ratio of a H 2O 2 flux (given in μM/min - 1) to catalase concentration (given in μM) of 10 min - 1 and above, H 2O 2 degradation occurred via the catalatic cycle. At lower ratios, however, H 2O 2 degradation proceeded with increasingly diminished production of O 2. At a ratio of 1 min - 1, O 2 formation could no longer be observed, although the enzyme still degraded H 2O 2. These results strongly suggest that at low physiological H 2O 2 fluxes H 2O 2 is preferentially metabolised reductively to H 2O, without release of O 2. The pathways involved in the reductive metabolism of H 2O 2 are presumably those previously reported as inactivation and reactivation pathways. They start from compound I and are operative at low and high H 2O 2 fluxes but kinetically outcompete the reaction of compound I with H 2O 2 at low H 2O 2 production rates. In the absence of NADPH, the reducing equivalents for the reductive metabolism of H 2O 2 are most likely provided by the protein moiety of the enzyme. In the presence of NADPH, they are at least in part provided by the coenzyme. 相似文献
12.
There is increasing evidence that hydrogen peroxide (H 2O 2) may act as a neuromodulator in the brain, as well as contributing to neurodegeneration in diseased states, such as Parkinson's disease. The ability to monitor changes in endogenous H 2O 2 in vivo with high temporal resolution is essential in order to further elucidate the roles of H 2O 2 in the central nervous system. Here, we describe the in vitro characterization of an implantable catalase-based H 2O 2 biosensor. The biosensor comprises two amperometric electrodes, one with catalase immobilized on the surface and one without enzyme (blank). The analytical signal is then the difference between the two electrodes. The H 2O 2 sensitivity of various designs was compared, and ranged from 0 to 56 ± 4 mA cm −2 M −1. The most successful design incorporated a Nafion ® layer followed by a poly- o-phenylenediamine (PPD) polymer layer. Catalase was adsorbed onto the PPD layer and then cross-linked with glutaraldehyde. The ability of the biosensors to exclude interference from ascorbic acid, and other interference species found in vivo, was also tested. A variety of the catalase-based biosensor designs described here show promise for in vivo monitoring of endogenous H 2O 2 in the brain. 相似文献
13.
The toxicity of H 2O 2 in Escherichia coli wild type and superoxide dismutase mutants was investigated under different experimental conditions. Cells were either grown aerobically, and then treated in M9 salts or K medium, or grown anoxically, and then treated in K medium. Results have demonstrated that the wild type and superoxide dismutase mutants display a markedly different sensitivity to both modes of lethality produced by H 2O 2 (i.e. mode one killing, which is produced by concentrations of H 2O 2 lower than 5 mM, and mode two killing which results from the insult generated by concentrations of H 2O 2 higher than 10 mM). Although the data obtained do not clarify the molecular basis of H 2O 2 toxicity and/or do not explain the specific function of superoxide ions in H 2O 2-induced bacterial inactivation, they certainly demonstrate that the latter species plays a key role in both modes of H 2O 2 lethality. A mechanism of H 2O 2 toxicity in E. coli is proposed, involving the action of a hypothetical enzyme which should work as an O 2-• generating system. This enzyme should be active at low concentrations of H 2O 2 (<5 mM) and high concentrations of the oxidant (>5 mM) should inactivate the same enzyme. Superoxide ions would then be produced and result in mode one lethality. The resistance at intermediate H 2O 2 concentrations may be dependent on the inactivation of such enzyme with no superoxide ions being produced at levels of H 2O 2 in the range 5–10 mM. Mode two killing could be produced by the hydroxyl radical in concert with superoxide ions, chemically produced via the reaction of high concentrations of H 2O 2 (>10 mM) with hydroxyl radicals. The rate of hydroxyl radical production may be increased by the higher availability of Fe 2+ since superoxide ions may also reduce trivalent iron to the divalent form. 相似文献
14.
Tea ( Camellia sinensis) catechins have been studied for disease prevention. These compounds undergo oxidation and produce H 2O 2. We have previously shown that holding tea solution or chewing tea leaves generates high salivary catechin levels. Herein, we examined the generation of H 2O 2 in the oral cavity by green tea solution or leaves. Human volunteers holding green tea solution (0.1-0.6%) developed salivary H 2O 2 with C max = 2.9-9.6 μM and AUC 0 → ∞ = 8.5-285.3 μM min. Chewing 2 g green tea leaves produced higher levels of H 2O 2 (C max = 31.2 μM, AUC 0 → ∞ = 1290.9 μM min). Salivary H 2O 2 correlated with catechin levels and with predicted levels of H 2O 2 (C max(expected) = 36 μM vs C max(determined) = 31.2 μM). Salivary H 2O 2 and catechin concentrations were similar to those that are biologically active in vitro. Catechin-generated H 2O 2 may, therefore, have a role in disease prevention by green tea. 相似文献
15.
The Emerson-Trinder reaction has been optimized in this work using an initial rate spectrophotometric method and response surface methodology (RSM). In this investigation, the variation range of critical variables along with the fixed parameters were selected based on a preliminary 'one at a time' (OVAT) procedure for the subsequent RSM chemometric analysis as follows: pH (6-10), buffer concentration (50-250 mM), 4-aminoantipyrine (4-AAP) concentration (1-5 mM), temperature (25-45°C). The optimum values of fixed parameters were: 4-fluorophenol (4-FP, 30 mM), horseradish peroxidase (HRP) enzyme activity (0.12 U mL -1), and the fixed concentration of the H 2O 2 in the chemometric experiments was 11.4 µM. The non-linear nature of the experimental response of the reaction system was explained by a second-order polynomial equation, which revealed the impact of the experimental factors, their interactions and also their optimum values. The results of the reported RSM analysis proved to be quite appropriate for the design and optimization of this reaction, as illustrated by the relatively high value of the determination coefficient (R 2=96.7%) for the fitting of quadratic model, along with the satisfactory results generated by the analysis of variance (ANOVA). All the evaluated analytical characteristics of this method: typical reaction progress curves, resulting linear calibration curve, within-day precisions at low and at high levels, and the upper and lower detection limits were, also, reported. In addition, to check the quality of the optimization and validity of the model, the assay of H 2O 2, in pooled serum matrix and in cosmetic samples, was performed. 相似文献
16.
Manganese peroxidases (MnP) from Phanerochaete chrysosporium and Bjerkandera sp. BOS55 were immobilised in glutaraldehyde–agarose gels. Four different strategies were considered concerning the activation of the support (low or high density) and the ionic strength (low or high). In terms of immobilisation rate and yield, better results were obtained when low ionic strength conditions and high density activated support (75 μEq/ml) were used. Immobilisation proceeds initially with an ionic adsorption which facilitates the further covalent attachment of the enzyme to the support. An almost complete immobilisation has been attained in a very short period (0.5–2 h). Immobilisation maintained a high percentage of MnP activity for long periods of time (activity levels of 50–60% after more than 1 year at room temperature storage). Other desirable effects such as increased thermostability at 50–60 °C for MnP from Bjerkandera and higher resistance to high H 2O 2 concentrations for MnP for P. chrysosporium were also obtained. This latter is quite an interesting feature because it avoids the inactivation of the enzyme in the presence of an unbalanced concentration of H 2O 2. The improved characteristics of the immobilised MnP make its application in several fields such as the enzymatic oxidation of hardly degradable compounds more feasible. 相似文献
17.
We had earlier shown that higher concentration of hydrogen peroxide (H 2O 2) induced p53-dependent apoptosis in glioma cell line with wild type p53 but had minimal effect on cells with mutated p53. Here we show a potentiating effect of hydroxylamine (HA), an inhibitor of catalase, on a nontoxic dose of H 2O 2 in glioma cells. HA sensitized both p53 wild type and mutated glioma cells to 0.25 mM H 2O 2. Potentiating effect of HA was independent of p53. Higher levels of reactive oxygen species (ROS) generation were observed in cells treated with HA+H 2O 2 as compared to cells treated with each component alone in both the cell lines. Dimethyl sulfoxide (DMSO) protected cells. Cytosolic cytochrome c and activated caspase 3 were detected at 4 h. The results suggest that higher levels of intracellular ROS, generated by HA+H 2O 2 act as a molecular switch in activating a rapidly acting p53-independent mitochondrial apoptotic pathway. 相似文献
18.
This work presents a novel, miniature optical biosensor by immobilizing horseradish peroxidase (HRP) or the HRP/glucose oxidase (GOx) coupled enzyme pair on a CMOS photosensing chip with a detection area of 0.5 mm × 0.5 mm. A highly transparent TEOS/PDMS Ormosil is used to encapsulate and immobilize enzymes on the surface of the photosensor. Interestingly, HRP-catalyzed luminol luminescence can be detected in real time on optical H 2O 2 and glucose biosensors. The minimum reaction volume of the developed optical biosensors is 10 μL. Both optical H 2O 2 and glucose biosensors have an optimal operation temperature and pH of 20–25 °C and pH 8.4, respectively. The linear dynamic range of optical H 2O 2 and glucose biosensors is 0.05–20 mM H 2O 2 and 0.5–20 mM glucose, respectively. The miniature optical glucose biosensor also exhibits good reproducibility with a relative standard deviation of 4.3%. Additionally, ascorbic acid and uric acid, two major interfering substances in the serum during electrochemical analysis, cause only slight interference with the fabricated optical glucose biosensor. In conclusion, the CMOS-photodiode-based optical biosensors proposed herein have many advantages, such as a short detection time, a small sample volume requirement, high reproducibility and wide dynamic range. 相似文献
19.
The effect of a range of iron chelates on the cytotoxicity of H 2O 2 was studied on a mammalian epithelial cell line. Iron complexes which were internalised enhanced the cytotoxicity of H 2O 2 measured by delayed thymidine incorporation. Iron complexed to 8-hydroxyquinoline (Fe/8-HQ) potentiated the cytotoxicity of 50 µM by 38% and Fe/dextran by 23%. Pre-exposure of cells to Fe/dextran at 4°C did not result in any potentiation of H 2O 2-induced cytotoxicity which we ascribe to failure of the Fe/dextran to be endocytosed at low temperature. Iron complexes which are slowly taken up or remain extracellular protected the cells from H 2O 2-induced cytotoxicity. Thus, Fe/EDTA inhibited the cytotoxicity of 50 µM H 2O 2 by 33%; Fe/ADP by 80% and Fe/ATP by 88%, suggesting mutual extracellular detoxification. 相似文献
20.
Cordyceps militaris mycelium produced mainly Cu, Zn containing superoxide dismutase (Cu, Zn-SOD). Cu, Zn-SOD activity was detectable in the culture filtrates, and intracellular Cu, Zn-SOD activity as a proportion protein was highest in early log phase culture. The effects of Cu 2+, Zn 2+, Mn 2+ and Fe 2+ on enzyme biosynthesis were studied. The Cu, Zn-SOD was isolated and purified to homogeneity from C. militaris mycelium and partially characterized. The purification was performed through four steps: (NH 4) 2SO 4 precipitation, DEAE-sepharose™ fast flow anion-exchange chromatography, CM-650 cation-exchange chromatography, and Sephadex G-100 gel filtration chromatography. The purified enzyme had a molecular weight of 35070 ± 400 Da and consisted of two equal-sized subunits each having a Cu and Zn element. Isoelectric point value of 7.0 was obtained for the purified enzyme. The N-terminal amino acid sequence of the purified enzyme was determined for 12 amino acid residues and the sequences was compared with other Cu, Zn-SODs. The optimum pH of the purified enzyme was obtained to be 8.2–8.8. The purified enzyme remained stable at pH 5.8–9.8, 25 °C and up to 50 °C at pH 7.8 for 1.5 h incubation. The purified enzyme was sensitive to H 2O 2, KCN. 2.5 mM NaN 3, PMSF, Triton X-100, β-mercaptoethanol and DTT showed no significant inhibition effect on the purified enzyme within 5 h incubation period. 相似文献
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