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1.
2-haloacid dehalogenases constitute a group of dehalogenases which are capable of dehalogenating the halogenated organic compounds. So far, the 2-haloacid dehalogenases have been found in many bacteria, but not in Paracoccus genus. In the present study, one enzyme 2-haloacid dehalogenase(designated as Deh99), induced by DL-2-chloropropionate(DL-2-CPA), was purified from the marine bacterium Paracoccus sp. DEH99, isolated from marine sponge Hymeniacidon perlevis. The enzyme of Deh99 was purified to homogeneity by ammonium sulfate precipitation, ion exchange chromatography(Q-Sepharose HP), and Superdex 200 gel filtration chromatography. The molecular weight of Deh99 was estimated to be 25.0 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE), and 50.0 kDa natively by gel filtration chromatography. The enzyme of Deh99 stereospecifically dehalogenated L-2-CPA to produce D-lactate, with an apparent Michaelis-Menten constant(Km) value of 0.21 mmol L-1 for L-2-CPA. The optimal pH and temperature for Deh99 activity were 10.0 and 40℃, respectively. The enzyme of Deh99 acted on short-carbon-chain 2-haloacids, with the highest activity towards monochloroacetate. The activity of Deh99 was slightly affected by DTT and EDTA, but strongly inhibited by Cu2+ and Zn2+. The enzyme of Deh99 shows unique substrate specificity and inhibitor sensitivities compared to previously characterized 2-haloacid dehalogenases and is the reported one about purified 2-haloacid dehalogenase isolated from the bacteria of Paracoccus genus.  相似文献   

2.
A bacterium hydrolyzing carboxymethylcellulose, isolated from Antarctic sea ice, was identified as Pseudoalteromonas sp. based on 16S rDNA gene sequences and named as Pseudoalteromonas sp. AN545. The extracellular endo-1,4-β-glucanase AN-1 was purified successively by ammonium sulfate precipitation, DEAE-Sepharose ion exchange chromatography and Sephadex G-75 gel filtration chromatography. The molecular mass of AN-1 was estimated to be 47.5 kDa utilizing SDS-PAGE and gel chromatography analysis. AN-1 could hydrolyze caboxymethylcellulose, avicel and β-glucan, but not cellobiose, xylan and p-Nitrophenyl-β-D-glucopyranoside. The optimal temperature and pH for the β-glucanase activity of AN-1 were determined to be at 30°C and pH 6.0, respectively. AN-1 was stable at acidic solutions of pH 5.0-6.5 and temperatures below 30°C for 1 h. Moreover, the specific activity was enhanced by Ca2+ and Mg2+, and inhibited by Cu2+. The kinetic parameters Michaelis constant (Km) and maximum velocity (Vmax) of AN-1 were 3.96 mg/mL and 6.06×10-2 mg/(min·mL), respectively.  相似文献   

3.
An oligopeptide permease A(OppA)was purified from the extracellular product of Vibrio harveyi SF-1.The molecular weight of the purified protein was estimated to be 58 kDa on SDS-PAGE.The purified protein showed phospholipase C activity at the optimal values of temperature 50℃ and pH 8.0.The enzymatic activity decreased when the temperature increased to 40℃.The N-terminal sequence of the purified protein was determined as ADVPAGTKLA,which is similar to that of OppA.The OppA pre-cursor gene was cloned from th...  相似文献   

4.
An alkaline protease from Acetes chinensis was purified and characterized in this study. The steps of purification include ammonium sulfate precipitation, ion-exchange chromatography with Q-sepharose Fast Flow, gel filtration chromatography with S300 and the second ion-exchange chromatography with Q-sepharose Fast Flow. The protease was isolated and purified, which was present and active on protein substrates (azocasein and casein). The specific protease activity was 17.15 folds and the recovery was 4.67. The molecular weight of the protease was estimated at 23.2 kD by SDS-PAGE. With azocasein as the susbstrate, the optimal temperature was 55°C and the optimal pH value was 5.5. Ion Ca2+ could enhance the proteolytic activity of the protease, while Cu2+, EDTA and PMSF could inhibit its activity.  相似文献   

5.
We isolated a bacterial strain (HC4) that is able to degrade κ-carrageenan from a live specimen of the red alga Hyalosiphonia caespitosa. With 16S rRNA gene sequencing, we identified the strain as Tamlana sp., and then purified an extracellular κ-carrageenase from a culture of Tamlana sp. HC4 by ammonium sulfate precipitation, Sephadex G-200 gel filtration chromatography, and DE-cellulose 52 anion-exchange chromatography. The purified enzyme yields a single band on SDS-PAGE with a molecular mass of 66.4 kDa. The optimal pH and temperature for κ-carrageenase activity are at 8.0 and 30°C, respectively. The enzyme is stable over the range of pH 7.2–8.6 below 45°C. The enzyme activity is strongly inhibited by Zn2+ and Cu2+ at 1 mmol/L. The enzyme-catalyzed reaction follows Michaelis-Menten kinetics with the Michaelis constant (K m ) at 7.63 mg/ml. Analysis of the degradation products of the κ-carrageenase by ESI-MS and 13C-NMR spectroscopy indicates that the enzyme degrades κ-carrageenan down to the level of κ-neocarrabiose sulfate.  相似文献   

6.
A lipase gene, lip1233, isolated from Pseudoalteromonas lipolytica SCSIO 04301, was cloned and expressed in E. coli. The enzyme comprised 810 amino acid residues with a deduced molecular weight of 80 kDa. Lip1233 was grouped into the lipase family X because it contained a highly conserved motif GHSLG. The recombinant enzyme was purified with Ni-NTA affinity chromatography. The optimal temperature and pH value of Lip1233 were 45°C and 8.0, respectively. It retained more than 70% of original activity after being incubated in pH ranging from 6.0 to 9.5 for 30 min. It was stable when the temperature was below 45°C, but was unstable when the temperature was above 55°C. Most metal ions tested had no significant effect on the activity of Lip1233. Lip1233 remained more than original activity in some organic solvents at the concentration of 30% (v/v). It retained more than 30% activity after incubated in pure organic solvents for 12 h, while in hexane the activity was nearly 100%. Additionally, Lip1233 exhibited typical halotolerant characteristic as it was active under 4M NaCl. Lip1233 powder could catalyze efficiently the synthesis of fructose esters in hexane at 40°C. These characteristics demonstrated that Lip1233 is applicable to elaborate food processing and organic synthesis.  相似文献   

7.
Macroalgae in a littoral zone are inevitably exposed to UV-B irradiance.We analyzed the effects of UV-B on isoenzyme patterns and activities of superoxide dismutase(SOD),peroxidase(POX),catalase(CAT),and ascorbate peroxidase(APX) of red algae Grateloupia filicina(Lamour.) C.Agardh.The activities of SOD,CAT,and APX changed in response to UV-B in a time- and dose-dependent manner.POX activity increased significantly under all three UV-B treatments.The enzymatic assay showed three distinct bands of SODⅠ(Mn-SOD),SODⅡ(Fe-SOD),and SODⅢ(CuZn-SOD) under a low(Luv) and medium(Muv) dose of UV-B irradiation,while SODⅠ and SODⅢ activities decreased significantly when exposed to a high dose of UV-B irradiation(Huv).The activity of POX isoenzymes increased significantly after exposure to UV-B,which is consistent with the total activity.In addition,a clear decrease in activity of CATIV was detected in response to all the three doses of UV treatments.Some bands of APX isoenzyme were also clearly influenced by UV-B irradiation.Correspondingly,the daily growth rate declined under all the three exposure doses,and was especially significant under Muv and Huv treatments.These data suggest that,although the protection mechanisms of antioxidant defense system are partly inducible by UV-B to prevent the damage,G.filicina has incomplete tolerance to higher UV-B irradiation stress.  相似文献   

8.
A new extracellular κ-carrageenase, namely CgkP, 34.0 kDa in molecular weight, was purified from Pseudoalteromonas sp. QY203. CgkP showed relatively high activity at acidities ranging from pH6.0 to pH9.0 and temperatures ranging from 30℃ to 50℃ with the highest activity at 45℃ and pH7.2. Sodium chloride increased its activity markedly, and KCl increased its activity slightly. The divalent and trivalent metal ions including Cu2+ , Ni2+ , Zn2+ , Mn2+ , Al3+ and Fe3+ significantly inhibited its activity, while Mg2+ did not. CgkP remained 70% of original activity after being incubated at 40℃ for 48 h, and remained 80% of the activity after being incubated at 45℃ for 1 h. It exhibited endo-κ-carrageenase activity, mainly depolymerizing the κ-carrageenan into disaccharide and tetrasaccharide. CgkP was more thermostable than most of previously reported κ-carrageenases with a potential of being used in industry.  相似文献   

9.
Activities of hexokinase （HK）, pyruvate kinase （PK）, superoxide dismutase （SOD） and catalase （CAT） and Hsp70 level were measured to evaluate the response of the commercially important sea cucumber （Apostichopus japonicus Selenka） to rapid temperature changes in laboratory. Animals were subjected to a higher temperature （from 10 to 20℃） （Tinc treatment） or to a lower temperature （from 20 to 10℃） （Tddec treatment） for 72 h. At 1, 3, 12, 24, 72 h of exposure, animals were removed and prepared for further analysis. Results showed that the effect of acute temperature changes on enzyme activities was significant. In Tinc treatment, activities of SOD and CAT increased immediately. The significant enhancement of SOD and CAT activities suggested that oxidative stress increases significantly when ambient temperature increasing from 10 to 20℃. The up-regulation of Flsp70 in Tinc and Tdec treatments indicated that Hsp70 was a bioindicator of thermal stress in the sea cucumber, and the expression pattern depended on the thermal treatment.  相似文献   

10.
Protease is wildly used in various fields, such as food, medicine, washing, leather, cosmetics and other industrial fields. In this study, an alkaline protease secreted by Micrococcus NH54PC02 isolated from the South China Sea was purified and characterized. The growth curve and enzyme activity curve indicated that the cell reached a maximum concentration at the 30th hour and the enzyme activity reached the maximum value at the 36th hour. The protease was purified with 3 steps involving ammonium sulfate precipitation, ion-exchange chromatography and hydrophobic chromatography with 8.22-fold increase in specific activity and 23.68% increase in the recovery. The molecular mass of the protease was estimated to be 25 kDa by SDS-PAGE analysis. The optimum temperature and pH for the protease activity were 50°C and pH 10.0, respectively. The protease showed a strong stability in a wide range of pH values ranging from 6.0–11.0, and maintained 90% enzyme activity in strong alkaline environment with pH 11.0. Inhibitor trials indicated that the protease might be serine protease. But it also possessed the characteristic of metalloprotease as it could be strongly inhibited by EDTA and strongly stimulated by Mn2+. Evaluation of matrix-assisted laser desorption ionization/time-of-flight MS (MALDI-TOF-TOF/MS) showed that the protease might belong to the peptidase S8 family.  相似文献   

11.
Jiang  Weiwei  Du  Meirong  Fang  Jianguang  Gao  Yaping  Mao  Yuze  Chen  Qionglin  Lin  Fan  Jiang  Zengjie 《中国海洋湖沼学报》2019,37(1):321-329

Water temperature is generally considered to be a major factor affecting the physiological and biochemical activities of marine bivalves. Here, the physiological and biochemical responses of Yesso scallop, Patinopecten yessoensis, to acute water temperature changes in summer were studied. Scallops were transferred directly to a lower temperature (Tdec treatment) (from 23°C to 15°C) or to a higher temperature (Tdec treatment) (from 15°C to 23°C) for 72 h, respectively. Results showed that the oxygen consumption and ammonia-N excretion rates of P. yessoensis decreased significantly in the Tdec treatment but increased dramatically at 6 h in the Tdec treatment (P <0.05). In the T dec treatment, hepatopancreas antioxidant enzyme activities, superoxide dismutase (SOD) and catalase (CAT) activities, increased substantially within 72 h (P <0.05). However, a significant decrease in CAT activity was found at 12 h in the Tdec treatment (P <0.01). A significant enhancement of acid phosphatase (ACP) activity and malondialdehyde (MDA) content was detected when scallops were acutely exposed to a temperature of 15°C. The levels of Cu/Zn-SOD gene expression in their gills up-regulated significantly in response to acute temperature changes (P <0.01). These data suggest that acute temperature change affects physiological and biochemical functions, and improve our knowledge of P. yessoensis under conditions of thermal stress.

  相似文献   

12.
The clotting protein (CP) plays important and diverse roles in crustaceans, such as coagulation and lipid transportation. A clotting protein was purified from the hemolymph of Chinese shrimp Fenneropenaeus chinensis (named as Fc-CP) with Q sepharose HP anion-exchange chromatography and phenyl sepharose HP hydrophobic interaction chromatography. Fc-CP was able to form stable clots in vitro in the presence of hemocyte lysate and Ca2+, suggesting that the clotting reaction is catalyzed by a Ca2+-dependent transglutaminase in shrimp hemocytes. The molecular mass of Fc-CP was 380 kDa under non-reducing conditions and 190 kDa under reducing conditions as was determined with SDS-PAGE. CP exists as disulfide-linked homodimers and oligomers. The N-terminal amino acid sequence of Fc-CP was identical to that of shrimps including Penaeus monodon, Farfantepenaeus paulensis and Litopenaeus vannamei; and similar to that of other decapods. The purified Fc-CP was digested with trypsin and verified on an ABI 4700 matrix-assisted laser desorption/ionization tandem time-of-flight (MALDI-TOF/TOF) mass spectrometry. Our results will aid to better understanding the coagulation mechanism of shrimp hemolymph.  相似文献   

13.
The growth, metabolism and physiological response of the sea cucumber, Apostichopus japonicus, were investigated during periods of inactivity. The body weight, oxygen consumption rate (OCR), activities of acidic phosphatase (ACP), alkaline phosphatase (AKP), catalase (CAT) and superoxide dismutase (SOD), and content of heat shock protein 70 (Hsp70) in the body wall and coelomic fluid of A. japonicus were measured during starvation, experimental aestivation and aestivation. The results showed that the body weight of sea cucumber in the three treatments decreased significantly during the experimental period (P < 0.05). The OCR of sea cucumber reduced in starvation and experimental aestivation treatments, but increased gradually in natural aestivation treatment. The activities of ACP and AKP of sea cucumber decreased gradually in all treatments, whereas those of SOD and CAT as well as Hsp70 content decreased in the starvation and experimental aestivation treatments and increased in natural aestivation treatment. The sea cucumber entered a state of aestivation at 24℃. To some extent, the animals in experimental aestivation were different from those in natural aestivation in metabolism and physiological response. These findings suggested that the aestivation mechanism of A. japonicus is complex and may not be attributed to the elevated temperature only.  相似文献   

14.
Ontogeny of the immune system in rock bream Oplegnathus fasciatus   总被引：1，自引：1，他引：0  
Histogenesis of the immune system and specific activity of superoxide dismutase(SOD) were studied in rock bream Oplegnathus fasciatus from fertilization to 50 days after hatching(DAH).The pronephric tubule primordium developed in the embryo,14 h 30 min post fertilization.The spleen anlage was observed between the swim bladder and the intestine at 5 DAH,and the thymus was formed as a paired structure under the pharyngeal epithelium above the gill arch at 10 DAH.The order of the immune organs becoming lymphoid was the pronephric kidney(10 DAH),thymus(15 DAH) and spleen(21 DAH).As the embryo developed,the specific activity of SOD gradually increased until hatching,but subsequently SOD activity continuously decreased to a minimum at 14 DAH.After the spleen became lymphoid,the specific activity of SOD was relatively stable.It is suggested that the immaturity of the lymphoid organs and low specific activity of SOD was the cause of the high mortality of fingerlings 12 to 16 DAH.  相似文献   

15.
The typical organic pollutant polycyclic aromatic hydrocarbon (PAH) anthracene was selected as a contaminant to investigate its effects on the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) in the clam Ruditapes philippinarum. The results show that SOD, CAT and GSH-Px had diff erent induction and inhibition reactions to anthracene stress, and that three diff erent organs in R. philippinarum (visceral mass, muscle tissue and mantle) had diff erent sensitivities to anthracene stress. This study suggest that SOD activities of the visceral mass, CAT activitities of the mantle and the visceral mass, and GSH-Px activity of the muscle tissue could be used as sensitive indicators of anthracene stress in R. philippinarum.  相似文献   

16.
Natural populations ofG. asiatica Zhang & Xia andG. lemaneiformis (Bory) Weber van Bosse were studied during 1984 and 1986 in Zhanshan Bay, Qingdao (36°4'N, 120° 21'E). Rapid growth (length, weight) of these plants occurred between mid-May and late June (water temperatures, 15–20°C). The major epiphyte ofG. asiatica wasEnteromorpha linza, whilePunctaria latifolia was the major epiphyte ofG. lemaneiformis. Epiphytism declined throughout early summer, and epiphytes were rare after mid-July (1984); they did not reappear in late summer, although macrophyte growth declined abruptly after early July. Populations ofG. asiatica varied during late spring-early summer between adjacent sandy and rocky portions of the intertidal zone; plants at the sandy site were larger and epiphyte-free. Amphipod densities were low on both species ofGracilaria, but the most abundant species wereAmpithoe lacertosa, Caprella equilibra, C. krøyeri, C. scauraandPontogeneia rostrata. Additional information on general community structure is provided for theG. asiatica zone.  相似文献   

17.
Uniconazole, as a plant growth retardant, can enhance stress tolerance in plants, possibly because of improved antioxidation defense mechanisms with higher activities of superoxide dismutase(SOD) and peroxidase(POD) enzymes that retard lipid peroxidation and membrane deterioration. These years much attention has been focused on the responses of antioxidant system in plants to uniconazole stress, but such studies on aquatic organism are very few. Moreover, no information is available on growth and antioxidant response in marine microalgae to uniconazole. In this paper, the growth and antioxidant responses of two marine microalgal species, Platymonas helgolandica and Pavlova viridis, at six uniconazole concentrations(0-15 mg L-1) were investigated. The results demonstrated that 3 mg L-1 uniconazole could increase significantly chlorophyll a and carbohydrate contents of P. helgolandica(P 0.05). Higher concentrations(≥12 mg L-1) of uniconazole could inhibit significantly the growth, dry weight, chlorophyll-a and carbohydrate contents of P. helgolandica and P. viridis(P 0.05). Uniconazole caused a significant increase in lipid peroxidation production(MDA) at higher concentrations(≥ 9 mg L-1). The activities of antioxidant enzymes, superoxide dismutase(SOD) and catalase(CAT) were enhanced remarkably at low concentrations of uniconazole. However, significant reduction of SOD and CAT activities was observed at higher concentrations of uniconazole.  相似文献   

18.
The Fe protein ofAnabaena cylindrica was first separated and purified by chromatography through DEAE-cellulose columns then by gel electrophoresis. The specific activity was up to 142.46 nmol C2H4/mg protein · min. It was homogeneous as shown by 1) a single band in the gel electrophorogram; 2) absence of Mo and tryptophan; 3) content of about 3.4 atoms of Fe per mole protein. The molecular weight of the Fe protein ofA. cylindrica was about 61,000 daltons as estimated by SDS-gel electrophoresis and calculated from the amino acid composition. The residues of aspartate and glutamate were about 2.6 times that of arginine and lysine in the Fe protein. Crossing Fe protein ofA. cylindrica with Mo?Fe protein ofAzotobacter vinelandii gave positive result. The reciprocal crossing also showed activity.  相似文献   

19.
In the present research, the strain SLYY-3 was isolated from sediments of Jiaozhou Bay, Qingdao, China. The strain SLYY-3, which produced a bacteriocin-like substance(BLS), was characterized to be a strain of Bacillus subtillis by biochemical profiling and 16 S r DNA sequence analysis. It is the first time to report that Bacillus subtilis from Jiaozhou Bay sediments could produce a BLS. The BLS of B. subtillis SLYY-3 exhibited strong inhibitory activity against gram-positive bacteria(including Staphylococcus aureus and B. subtillis) and some fungi(including Penicillium glaucum, Aspergillus niger and Aspergillus flavus). The antimicrobial activity was detected from culture in the exponential growth phase and reached its maximum when culture entered into stationary growth phase. It was thermo-tolerant even when being kept at 100℃ for 60 min without losing any activity and stable over a wide p H range from 1.0 to 12.0 while being inactivated by proteolytic enzyme and trypsin, indicating the proteinaceous nature of the BLS. The BLS was purified by precipitation with hydrochloric acid(HCl) and gel filteration(Sephadex G-100). SDS-PAGE analysis of the extracellular peptides of SLYY-3 revealed a bacteriocin-like protein with a molecular mass of 66 k Da. Altogether, these characteristics indicate the potential of the BLS for food industry as a protection against pathogenic and spoilage microorganisms.  相似文献   

20.
Low molecular weight peptide (LMWP) was prepared from clam Paphia undulate and its antiaging effect on D-galactose-induced acute aging in rats, aged Kunming mice, ultraviolet-exposed rats, and thermally injured rats was investigated. P. undulate flesh was homogenized and digested using papain under optimal conditions, then subjected to Sephadex G-25 chromatography to isolate the LMWP. Administration of LMWP significantly reversed D-galactose-induced oxidative stress by increasing the activities of glutathione peroxidase (GPx) and catalase (CAT), and by decreasing the level of malondialdehyde (MDA). This process was accompanied by increased collagen synthesis. The LMWP prevented photoaging and promoted dermis recovery and remission of elastic fiber hyperplasia. Furthermore, treatment with the LMWP helped to regenerate elastic fibers and the collagen network, increased superoxide dismutase (SOD) in the serum and significantly decreased MDA. Thermal scald-induced inflammation and edema were also relieved by the LWMP, while wound healing in skin was promoted. These results suggest that the LMWP from P. undulate could serve as a new antiaging substance in cosmetics.  相似文献   

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