Improvement of activity and stability of chloroperoxidase by chemical modification

<p>Abstract</p> <p>Background</p> <p>Enzymes show relative instability in solvents or at elevated temperature and lower activity in organic solvent than in water. These limit the industrial applications of enzymes.</p> <p>Results</p> <p>In order to improve the activity and stability of chloroperoxidase, chloroperoxidase was modified by citraconic anhydride, maleic anhydride or phthalic anhydride. The catalytic activities, thermostabilities and organic solvent tolerances of native and modified enzymes were compared. In aqueous buffer, modified chloroperoxidases showed similar <it>K</it>_mvalues and greater catalytic efficiencies <it>k</it>_cat/<it>K</it>_mfor both sulfoxidation and oxidation of phenol compared to native chloroperoxidase. Of these modified chloroperoxidases, citraconic anhydride-modified chloroperoxidase showed the greatest catalytic efficiency in aqueous buffer. These modifications of chloroperoxidase increased their catalytic efficiencies for sulfoxidation by 12%~26% and catalytic efficiencies for phenol oxidation by 7%~53% in aqueous buffer. However, in organic solvent (DMF), modified chloroperoxidases had lower <it>K</it>_mvalues and higher catalytic efficiencies <it>k</it>_cat/<it>K</it>_mthan native chloroperoxidase. These modifications also improved their thermostabilities by 1~2-fold and solvent tolerances of DMF. CD studies show that these modifications did not change the secondary structure of chloroperoxidase. Fluorescence spectra proved that these modifications changed the environment of tryptophan.</p> <p>Conclusion</p> <p>Chemical modification of epsilon-amino groups of lysine residues of chloroperoxidase using citraconic anhydride, maleic anhydride or phthalic anhydride is a simple and powerful method to enhance catalytic properties of enzyme. The improvements of the activity and stability of chloroperoxidase are related to side chain reorientations of aromatics upon both modifications.</p

Synergetic Effect of Yihuo Qingyi Decoction () and Recombinant Staphylokinase in Treatment of Severe Acute Pancreatitis of Rats

ObjectiveTo investigate the effect of recombinant staphylokinase (r-Sak) and the Chinese medicine Yihuo Qingyi Decoction (Herbal decoction for severe acute pancreatitis) in the treatment of the severe acute pancreatitis (SAP) in rats, and to observe the synergistic effect of the two.MethodsOne hundred and sixty-two adult male SD rats with the body mass of 250–280 g were randomly divided into the following 5 groups: sham operation group (n=18), control group (n=36), Yihuo Qingyi Decoction treatment group (n=36), r-Sak treatment group (n=36), and Yihuo Qingyi Decoction plus r-Sak treatment group (n=36). The SAP ratmodel was prepared by retrograde injection of 5% sodium taurocholate into the cholangiopancreatic duct. Two days before modeling, Yihuo Qingyi Decoction was intragastrically administrated, and r-Sak was intraperitoneally injected. The survival rate within 18 h after modeling was determined. The pancreatic blood flow, the weight of ascites, and the serum amylase and lipase were investigated at 6 h, 12 h, and 18kh after modeling, and the pancreatic tissue was examined under light microscopy to see its pathological change.ResultsThe 18 h survival count of group A,B,C,D and E rats was 9,2,6,7 and 8 respectively. After r-Sak and Yihuo Qingyi Decoction intervention, the serum amylase and lipase and the weight of ascites were significantly decreased, especially in group E.18 h after modeling, the level of the serum amylase and lipase and the weight of ascites in group E was 1 100±118 U·L−1,1 000±150 U·L−1 and 13.40±1.80 g respectively, obviously lower than that of group B (P<0.05). After SAP was induced, the pancreatic blood flow showed a tendency to decrease, but the decrease extent in the treatment groups was smaller than that in the control group.18h after modeling, the pancreatic blood flow in group B and group E was 30.16±8.96 mL·100 g−1·min−1, and 129.10±42.58 mL·100 g−1·min−1 respectively, there was significant difference (P<0.05). The pathological change of the pancreatic tissue was alleviated in the treatment groups.ConclusionBoth r-Sak and Yihuo Qingyi Decoction play a beneficial role in the treatment of rat SAP and there is a synergistic effect between the two

Structure and Properties of Dense Silica Glass

The O K-edge x-ray Raman scattering (XRS), Brillouin scattering and diffraction studies on silica glass at high pressure have been elucidated in a unified manner using model structures obtained from First-Principles molecular dynamics calculations. This study provides a comprehensive understanding on how the structure is related to the physical and electronic properties. The origin of the "two peak" pattern in the XRS is found to be the result of increased packing of oxygen near the Si and is not a specific sign for sixfold coordination. The compression mechanism involving the presence of 5- and 6-fold coordinated silicon is confirmed. A slight increase in the silicon-oxygen coordination higher than six was found to accompany the increase in the acoustic wave velocity near 140 GPa

Inhibitory effects of total saponins from Ilex pubescens Hook against hydrogen peroxide-induced cardiomyocyte apoptosis

Purpose: To study the protective effects of total saponins from Ilex pubescens Hook (IPTS) against cardiomyocyte apoptosis.Methods: Response surface methodology (RSM) based on Box-Benhnken Design (BBD) was carried out to optimize the extraction of IPTS. Thereafter, H9c2 cell model prepared by hydrogen peroxide (H2O2) treatment was used to investigate the effects of IPTS on cardiomyocyte apoptosis. Cell viability was determined using MTT assay, while the levels of lactate dehydrogenase (LDH), malondialdehyde (MDA), superoxide dismutase (SOD), creatine kinase (CK) and catalase (CAT) were measured as indices of oxidative stress. Expressions of proteins related to apoptosis (caspase-3, Bax and Bcl-2) were measured using Western blot assay.Results: Optimal IPTS extraction was achieved with extraction temperature of 86.6 °C, extraction time of 2.23 h and water: raw material ratio of 10.8 mL/g. IPTS extract, at doses of 200, 400, 600 and 800 μg/mL, significantly increased the viability of H2O2-treated H9c2 cells (p &lt; 0.05), but significantly decreased LDH and CK activities (p &lt; 0.01). It also led to significant increases in SOD and CAT activities, and significant decreases in the levels of MDA in these cells (p &lt; 0.01). There were significant down-regulation of the protein expressions of caspase-3 and Bax (p &lt; 0.01) in IPTS-treated H9c2 cells, as well as significant up-regulation of Bcl-2 protein expression (p &lt; 0.01).Conclusion: These results suggest that IPTS can protect cardiomyocytes against apoptosis via the inhibition of oxidative stress and mitochondria-induced intrinsic apoptosis.Keywords: Ilex pubescens, Total saponins, Cardiomyocytes, Apoptosis, H9c2 cell

Pinoresinol diglucoside exhibits protective effect on dexamethasone-induced osteoporosis in rats

Purpose: To investigate the effect of pinoresinol diglucoside (PDG) on dexamethasone-induced osteoporosis in rats.Methods: Sixty Wistar rats were randomly and equally divided into normal, control, alendronate and PDG (10, 20 or 40 mg/kg) groups. Bone tissue parameters, including length, transverse diameter, weight, bone mineral content (BMC) and bone mineral density (BMD), were determined using vernier caliper, electronic balance and single photon bone mineral density meter. Serum biochemical indices, including Ca2+, inorganic phosphorus (IP), IL-6, TNF-α and alkaline phosphatase (ALP), were determined using colorimetry and enzyme-linked immunosorbent assay (ELISA). Osteoprotegerin (OPG) and receptor activator of nuclear factor-κB ligand (RANKL) proteins were detected by Western blot.Results: PDG (10, 20 or 40 mg/kg) increased significantly (p &lt; 0.05 or 0.01) transverse diameter (3.64 – 3.79 vs. 3.31 mm), weight (0.73 – 0.78 vs. 0.67 g), BMC (0.16 – 0.23 vs. 0.12 g/cm), BMD (0.27 – 0.35 vs. 0.22 g/cm2) of right femur, serum Ca2+ level (2.16 – 2.39 vs. 1.94 mmol/L), and OPG level of left femur, compared with those in the control group. PDG (10, 20 or 40 mg/kg) reduced significantly (p &lt; 0.05 or 0.01) serum IP (1.34 – 1.14 vs. 1.76 mmol/L), IL-6 (103.25 – 95.38 vs. 108.74 ng/L), TNF-α (87.46 – 82.05 vs. 92.38 ng/L), ALP (334.79 – 276.32 vs. 486.45 U/L) levels or activities, and RANKL level of left femur, compared with those in the control group.Conclusion: PDG exhibits a protective effect on dexamethasone-induced osteoporosis by increasing bone mass and regulating bone metabolism. Thus, PDG may be a candidate drug for treating osteoporosis.Keywords: Pinoresinol diglucoside, Osteoporosis, Bone mass, Bone metabolism, Dexamethasone, Osteoprotegeri

Angiotensin-converting enzyme gene 2350 G/A polymorphism and susceptibility to atrial fibrillation in Han Chinese patients with essential hypertension

OBJECTIVE: The angiotensin-converting enzyme gene is one of the most studied candidate genes related to atrial fibrillation. Among the polymorphisms of the angiotensin-converting enzyme gene, the 2350 G/A polymorphism (rs4343) is known to have the most significant effects on the plasma angiotensin-converting enzyme concentration. The aim of the present study was to investigate the association of the angiotensin-converting enzyme 2350 G/A polymorphism with atrial fibrillation in Han Chinese patients with essential hypertension. METHODS: A total of 169 hypertensive patients were eligible for this study. Patients with atrial fibrillation (n = 75) were allocated to the atrial fibrillation group, and 94 subjects without atrial fibrillation were allocated to the control group. The PCR-based restriction fragment length polymorphism technique was used to assess the genotype frequencies. RESULTS: The distributions of the angiotensin-converting enzyme 2350 G/A genotypes (GG, GA, and AA, respectively) were 40.43%, 41.49%, and 18.08% in the controls and 18.67%, 46.67%, and 34.66% in the atrial fibrillation subjects (p = 0.037). The frequency of the A allele in the atrial fibrillation group was significantly greater than in the control group (58.00% vs. 38.83%, p = 0.0007). Compared with the wild-type GG genotype, the GA and AA genotypes had an increased risk for atrial fibrillation. Additionally, atrial fibrillation patients with the AA genotype had greater left atrial dimensions than the patients with the GG or GA genotypes (

Polysaccharides from the Medicinal Mushroom Cordyceps taii Show Antioxidant and Immunoenhancing Activities in a D-Galactose-Induced Aging Mouse Model

Cordyceps taii, an edible medicinal mushroom native to south China, is recognized as an unparalleled resource of healthy foods and drug discovery. In the present study, the antioxidant pharmacological properties of C. taii were systematically investigated. In vitro assays revealed the scavenging activities of the aqueous extract and polysaccharides of C. taii against various free radicals, that is, 1,1-diphenyl-2-picrylhydrazyl radical, hydroxyl radical, and superoxide anion radical. The EC50 values for superoxide anion-free radical ranged from 2.04 mg/mL to 2.49 mg/mL, which was at least 2.6-fold stronger than that of antioxidant thiourea. The polysaccharides also significantly enhanced the antioxidant enzyme activities (superoxide dismutase, catalase, and glutathione peroxidase) and markedly decreased the malondialdehyde production of lipid peroxidation in a D-galactose-induced aging mouse model. Interestingly, the immune function of the administration group was significantly boosted compared with the D-galactose-induced aging model group. Therefore, the C. taii polysaccharides possessed potent antioxidant activity closely associated with immune function enhancement and free radical scavenging. These findings suggest that the polysaccharides are a promising source of natural antioxidants and antiaging drugs. Consequently, a preliminary chemical investigation was performed using gas chromatography-mass spectroscopy and revealed that the polysaccharides studied were mainly composed of glucose, mannose, and galactose. Fourier-transform infrared spectra also showed characteristic polysaccharide absorption bands

Quantum Liouville Theory On The Riemann Sphere With n>3n>3 Punctures

We have studied the quantum Liouville theory on the Riemann sphere with n>3 punctures. While considering the theory on the Riemann surfaces with n=4 punctures, the quantum theory near an arbitrary but fixed puncture can be obtained via canonical quantization and an extra symmetry is explored. While considering more than four distinguished punctures, we have found the exchange relations of the monodromy parameters from which we can get a reasonable quantum theory.Comment: 13 page

{μ-trans-N,N′-Bis[(diphenyl­phosphan­yl)meth­yl]benzene-1,4-diamine-κ2 P:P′}bis­{(acetonitrile-κN)[dipyrido[3,2-a:2′,3′-c]phenazine-κ2 N 4,N 5]copper(I)} bis­(tetra­fluoridoborate)

In the centrosymmetric dinuclear title compound, [Cu2(C2H3N)2(C18H10N4)2(C32H30N2P2)](BF4)2, the CuI centre is coordinated by two N atoms from a dipyridophenazine ligand, one P atom from an N,N′-bis­[(diphenyl­phosphan­yl)meth­yl]benzene-1,4-diamine (bpbda) ligand, and one N atom from an acetonitrile mol­ecule in a distorted tetra­hedral geometry. The bpbda ligand, lying on an inversion center, bridges two CuI centres into a Z-shaped complex. Intra­molecular π–π inter­actions between the dipyridophenazine ligand and the benzene ring of the bpbda ligand are observed [centroid–centroid distance = 3.459 (3) Å]. The crystal structure also involves inter­molecular π–π inter­actions between the dipyridophenazine ligands [centroid–centroid distance = 3.506 (3) Å], which lead to a one-dimensional supra­molecular structure