共查询到20条相似文献,搜索用时 15 毫秒
1.
Takahashi S Obana Y Okada S Abe K Kera Y 《Journal of Bioscience and Bioengineering》2012,113(1):79-83
Tris(1,3-dichloro-2-propyl) phosphate (TDCPP), a flame retardant, is regarded as a potentially toxic and persistent environmental contaminant. We previously isolated a TDCPP-degrading bacterium, Sphingobium sp. strain TCM1, which, however, produced a toxic metabolite: 1,3-dichloro-2-propanol (1,3-DCP). This study was undertaken to develop a technique for complete TDCPP detoxification using strain TCM1 with a 1,3-DCP-degrading bacterium, Arthrobacter sp. strain PY1. For efficient detoxification, we designed a resting cell system and examined the effect of freezing and lyophilization treatments for preparation of their resting cells. Results show that treatments had no marked adverse effect on their activities. The TDCPP dephosphorylation by TCM1 resting cells was optimal at 30°C and pH 8.5. Also, 1,3-DCP dehalogenation by strain PY1 resting cells was optimal at 35°C and pH 9.5. Under those respective conditions, the activities were 2.48 μmol h− 1·OD660− 1 for TDCPP and 0.95 μmol h− 1·OD660− 1 for 1,3-DCP. Based on these results, we set the reaction temperature to 30°C and pH to 9.0. Then we examined the detoxification of 50 μM TDCPP using mixed resting cells at a final OD660 of 0.05 for strain TCM1 and 0.2 for strain PY1. In these conditions, TDCPP was eliminated after 1 h, but some of the resulting 1,3-DCP remained at a constant level. The increase in strain PY1 cells to a final OD660 of 4.0 decreased the TDCPP dephosphorylation rate of strain TCM1 cells but achieved complete detoxification of TDCPP during 12 h of reaction. 相似文献
2.
A novel serine protease, with a molecular mass of 19 kDa and the N-terminal sequence of ARTPEAPAEV, was isolated from dried fruiting bodies of the mushroom Pholiota nameko. The purification protocol comprised ion exchange chromatography on DEAE-cellulose, Q-Sepharose and SP-Sepharose, and gel filtration on Superdex 75. It was unadsorbed on DEAE-cellulose and Q-Sepharose but adsorbed on SP-Sepharose. It exhibited an optimum temperature at 50°C, an optimum pH at pH 8.8, a Km of 5.64 mg/mL and a Vmax of 0.98 μmol/min/mL against substrate casein. A number of metal ions inhibited the enzyme including Pb(2+), Mn(2+), Ca(2+), Hg(2+), Zn(2+), Cu(2+), Co(2+), Fe(3+) and Al(3+), with the inhibition of the last two cations being the most potent. K(+) and Mg(2+) slightly enhanced, while Li(+) moderately potentiated the activity of the protease. The protease was strongly inhibited by phenylmethylsulfonyl fluoride (PMSF), suggesting that it is a serine protease. 相似文献
3.
4.
Guoqing Zhang Hexiang Wang Xiaoqing Zhang TziBun Ng 《Journal of Bioscience and Bioengineering》2010,109(1):20-24
A 33.5-kDa serine protease designated as helvellisin was isolated from dried fruiting bodies of the wild ascomycete mushroom Helvella lacunosa. It was purified by using a procedure which entailed ion exchange chromatography on DEAE-cellulose, CM-Sepharose, Q-Sepharose, and FPLC-gel filtration on Superdex 75. The protease was characterized by unique N-terminal amino acid sequence, thermostability and pH stability. The protease exhibited a pH optimum of 11.0 and a temperature optimum of 65 °C, with about 40% activity remaining at 87 °C and pH 5 and 13. Helvellisin demonstrated a protease activity of 14 600 U/mg toward casein. The Km of the purified protease for casein was 3.81 mg/ml at pH 11.0 and 37 °C. The Vmax was 5.35 × 10− 2 mg ml− 1 min− 1. It was adversely affected by phenylmethylsulfonyl fluoride, suggesting that it is serine protease. The activity of the protease was enhanced by Mg2+, Fe2+ and Mn2+, but was curtailed by Cu2+, Hg2+ and Fe3+. It was devoid of antifungal and ribonuclease activities. 相似文献
5.
6.
Ye JH Wang LX Chen H Dong JJ Lu JL Zheng XQ Wu MY Liang YR 《Journal of Bioscience and Bioengineering》2011,111(2):232-236
An adsorption separation method using Polyamide-6 (PA) as an adsorbent was developed to separate catechins from green tea extract. The adsorption capacity of total catechins for PA was 193.128 mg g−1 with an adsorption selectivity coefficient KAB of total catechins over caffeine 21.717, which was better than macroporous resin model HPD 600. The Langmuir model and the pseudo-second order mode were primely fitted to describe its equilibrium data and adsorption kinetics, respectively. PA column separation by two-step elution using water and 80% (v/v) aqueous ethanol was established to prepare catechins complex which contained 670.808 mg g−1 total catechins and 1.828 mg g−1 caffeine. It is considered that PA was a promising adsorbent for selective isolation of catechins. 相似文献
7.
A rhizosphere strain of Stenotrophomonas maltophilia strain MUJ that is strongly antagonistic towards fungal phytopathogens secretes to the culture medium a single form of active chitinolytic enzyme belonging to family 18 of glycosyl hydrolases. The chitinase was purified by a two-stage procedure embracing fractionation with ammonium sulfate and affinity chromatography. The molecular mass of the purified enzyme determined by SDS-PAGE was approximately 52 kDa. The enzyme demonstrated highest activity at 45°C and pH 6.8. The enzymatic protein showed considerable thermal stability during 2 h incubation at 45°C. The activity of the enzyme was strongly inhibited in the presence of Hg2+ and Cu2+. By applying mass spectrometry analysis, the peptides derived from the purified chitinase were assigned to amino acid sequences of the type ChiA chitinases synthesized by Stenotrophomonas bacteria. The purified enzyme inhibited the growth of fungal phytopathogens belonging to the genera Fusarium, Rhizoctonia and Alternaria. 相似文献
8.
Natto-red bean with fibrinolytic activity was prepared by fermenting red beans with Bacillus subtilis. A fibrinolytic enzyme was purified from fermented natto-red bean by sequential steps of ammonium sulfate fractionation, Sephacryl S-200 HR gel filtration and PBE 94 chromatofocusing. Through these steps, the purity of the enzyme increased 291-fold with 1.5% activity recovery. SDS–PAGE and isoelectric focusing electrophoresis showed the molecular mass and pI of the purified enzyme to be 29.93 kDa and 6.35, respectively. When N-succinyl-Ala-Ala-Pro-Phe-ρNA was used as an enzyme substrate, the Km, Vmax, and optimal reaction pH and temperature were 0.59 mM, 79.4 μmole ρNA/min mg, 9 and 60 °C, respectively. Among the synthetic substrates, the most sensitive were N-succinyl-Ala-Ala-Pro-Phe-ρNA, followed by N-benzoyl-Val-Gly-Arg-ρNA. Chemical modifiers, such as phenylmethyl sulfonyfluoride, N-bromosuccinimide and N-ethyl-5-phenylisoxazolium-3′-sulfonate, almost completely inhibited the activity of the purified enzyme. These results indicated that the purified fibrinolytic enzyme was a subtilisin-like serine protease. 相似文献
9.
The textural properties of beef over the temperature range −18 to +5 °C were measured using Warner Bratzler (WB) and tensile techniques. In addition, the effects of rapid radio frequency (RF) tempering and slower conventional air tempering on texture were compared. Temperature showed a significant effect (P < 0.05) on WB and tensile shear force, with higher values obtained at temperatures on or below −5 °C. Work to fracture values showed two peaks at −15 and −3 °C. Sample thickness and muscle fibre direction were also important factors affecting shear force, with samples cut across fibres showing higher values. Tempering method showed no effect (P ? 0.05) on the textural properties measured. In light of the rapid nature of RF tempering, these findings will be of interest to the meat industry. 相似文献
10.
Our objective was to assess the effects of lactate enhancement in combination with different packaging systems on beef longissimus lumborum and psoas major steak color. Strip loins and tenderloins (n = 16) were assigned to one of four injection treatments (non-injected control, water-injected control, 1.25%, and 2.5% lactate in the finished product). Steaks were individually packaged in either vacuum, high-oxygen (80% O2/20% CO2), or 0.4% CO (30% CO2/69.6% N2) and stored for either 0, 5, or 9 days at 1 °C. The L∗ and a∗ values of both the longissimus and psoas responded similarly to lactate, which at 2.5% darkened steaks (P < 0.05) packaged in all atmospheres and improved (P < 0.05) the redness of steaks packaged in high-oxygen. Packaging steaks in CO did not counteract the darkening effects of lactate. Nevertheless, CO improved (P < 0.05) color stability compared with high-oxygen packaging. 相似文献
11.
An extracellular proteinase from Virgibacillus sp. SK33, isolated from 1 month-old fish sauce, was purified to electrophoretic homogeneity, using hydrophobic interaction chromatography and hydroxyapatite with purification fold of 2.5 and 7% yield. The anomalous molecular weight (MW) of 19 kDa was obtained from SDS–PAGE, whereas a MW of 33.7 kDa was determined by MALDI-TOF. Optimum conditions for catalytic activity were 55 °C and pH 7.5. The proteinase was strongly inhibited by phenylmethanesulfonyl fluoride (PMSF) and preferentially hydrolysed Suc-Ala-Ala-Pro-Phe-AMC, indicating a serine proteinase with subtilisin-like characteristics. Km and kcat of the purified proteinase were 27 μM and 12 s−1, respectively. Proteinase activity, toward both synthetic and anchovy substrates, increased with NaCl up to 25%. The proteinase exhibited high stability in both the absence and presence of NaCl up to 25%. Approximately 2.5-fold increase in activity was observed in the presence of divalent cations, including Ca2+, Mg2+ and Sr2+ at 100 mM. MALDI-TOF MS and LC–ESI-MS/MS analyses, as well as N-terminal sequences, revealed that the purified enzyme did not match microbial proteinases in the database, indicating it to be a novel proteinase. 相似文献
12.
The antifungal activity of Pimpinella anisum L. (anise), Pëumus boldus Mol (boldus), Hedeoma multiflora Benth (mountain thyme), Syzygium aromaticum L. (clove), and Lippia turbinate var. integrifolia (griseb) (poleo) essential oils (EOs) against Aspergillus section Flavi was evaluated in sterile maize grain under different water activity (aw) condition (0.982, 0.955, and 0.90). The effect of EOs added to maize grains on growth rate, lag phase, and aflatoxin B1 (AFB1) accumulation of Aspergillus section Flavi were evaluated at different water activity conditions. The five EOs analyzed have been shown to influence lag phase and growth rate. Their efficacy depended mainly on the essential oil concentrations and substrate water activity conditions. All EOs showed significant impact on AFB1 accumulation. This effect was closely dependent on the water activity, concentration, and incubation periods. Important reduction of AFB1 accumulation was observed in the majority of EO treatments at 11 days of incubation. Boldus, poleo, and mountain thyme EO completely inhibited AFB1 at 2000 and 3000 μg g−1. Inhibition of AFB1 accumulation was also observed when aflatoxigenic isolates grew with different concentration of EOs during 35 days. 相似文献
13.
Characterisation of esterase activities from the edible mushroom species, Amanita vaginata var. vaginata and Tricholoma terreum, were investigated. Native electrophoresis of the crude extracts prepared from both mushroom samples showed the presence of esterolytic activities. The extracts had the greatest activity in the presence of p-nitrophenyl butyrate (pNPB) as a substrate. pH and temperature optima were found to be 8.0 and 30 °C for both enzymes, respectively. Vmax and Km values were determined as 14.2 U/l and 71 μM for A. vaginata var. vaginata and 34.6 U/l and 9.6 μM for T. terreum, respectively. The pH-stability profile showed a stationary line between 3.0 and 10.0 for both enzymes. The esterolytic activities from the extracts were maintained between 10 and 40 °C for 4 h and started to decrease at 50 °C. The effects of EDTA, NaN3, DTT and PMSF on the enzyme activity were also investigated. 相似文献
14.
The presence of a protease in Artocarpus integer leaves, which are traditionally used as a meat tenderiser, was verified by the presence of a band at 69 kDa, using caseinolytic zymography. Purification by temperature phase partitioning with Triton X-114, ammonium sulphate precipitation and gel filtration chromatography yielded a preparation with a 12-fold increase in enzyme purity and a final specific activity of 76.67 U/mg. The cysteinic nature of this enzyme was confirmed through inhibition of enzyme activity by E-64 and iodoacetamide and enhancement of activity by cysteine and 2-mercaptoethanol. The protease retained 70% of its activity over a broad pH range (pH 6–12), with optimal activity recorded at pH 10 and 40 °C. The enzyme was stable at temperatures up to 70 °C, with 80% of its activity intact. Addition of 5 mM Ca2+ stimulated enzyme activity and a kinetic study of the enzyme yielded Km and Vmax values of 0.304 mg/mL and 0.735 mg/mL/min, respectively. 相似文献
15.
It is considered that consumption of very long chain (VLC, carbon chain length ?20) n − 3 PUFAs in most Western populations is sub-optimal and benefits in relation to chronic disease would be gained from increased consumption. This review examines the current contribution that meat makes to dietary intake of VLC n − 3 PUFA and given its current low contribution, how ruminant meat may be enriched. Enrichment both directly with VLC n − 3 fatty acids and indirectly by increasing intake by the animals of α-linolenic acid (ALNA; C18:3 n − 3) are considered. Since it now appears that dietary ALNA is a very limited source of VLC n − 3 PUFA in humans, the indirect route is controversial but since some forages are rich sources of ALNA this route has many sustainability and environmental attractions. Consideration is also given to the increased concentrations of trans and conjugated fatty acids that will arise from enriching ruminant meat with PUFA. 相似文献
16.
Inhibitory effects of 4-chlorosalicylic acid on mushroom tyrosinase and its antimicrobial activities 总被引:1,自引:0,他引:1
The inhibitory effects of 4-chlorosalicylic acid on the activity of mushroom tyrosinase have been investigated. The results showed that 4-chlorosalicylic acid could strongly inhibit both monophenolase activity and diphenolase activity. The IC50 values were estimated as 1.89 mM and 1.10 mM for monophenolase and diphenolase activities, respectively. For the monophenolase activity, 4-chlorosalicylic acid could not only lengthen the lag time, but also decrease the steady-state rate. For the diphenolase activity, kinetic analyses showed that the inhibition by 4-chlorosalicylic acid was reversible and its mechanism was mixed-II type, which is different from salicylic acid. The inhibition constants (KI and KIS) were determined to be 1.51 mM and 0.82 mM, respectively. Furthermore, the antibacterial activity against Escherichia coli, Bacillus subtilis and Staphyloccocus aureus and antifungal activity against Aspergillus niger and Candida boidinii were investigated. The results showed that 4-chlorosalicylic acid was the most effective against E. coli with the MIC of 250 μg/ml and with the MBC of 500 μg/ml. 相似文献
17.
The gluteus medius (GM) from USDA Select beef carcasses was used to test the effect of aging period on bloom development. Top sirloin butts (IMPS #184) were randomly allocated to 0, 7, 14, 21, 28, and 35 d vacuum-aging at 2 °C (n = 10/aging period). Each week, aged top sirloin butts were faced before two 2.5-cm-thick, non-adjacent steaks were cut and instrumental color (L∗, a∗, and b∗) of the GM was measured at 10-min intervals for 2 h after cutting. Steaks aged for 7 and 14 d were a more vivid (greater chroma values; P < 0.05), redder (greater a∗ values; P < 0.05), and more yellow (greater b∗ values; P < 0.05) color than steaks from the other aging periods. Change in total color (ΔE) was greater (P < 0.05) for steaks from top sirloin butts aged 7, 14, and 21 d than steaks from top butts aged 28 and 35 d, whereas oxymyoglobin percentages for steaks from top butts aged 7 and 14 days were greater (P < 0.05) than those from top sirloin butts aged 28 and 35 d. As much as 90% of the total increase (P < 0.05) in a∗, b∗, and chroma values, as well as hue angles and oxymyoglobin percentages, was achieved during the first 60 min after cutting. 相似文献
18.
Zhang D Spadaro D Valente S Garibaldi A Gullino ML 《International journal of food microbiology》2012,153(3):453-464
An alkaline protease gene was amplified from genomic DNA and cDNA of the antagonistic yeast-like fungus Aureobasidium pullulans PL5, a biocontrol agent effective against Monilinia laxa on stone fruit and Botrytis cinerea and Penicillium expansum on pome fruits. An open reading frame of 1248 bp encoding a 415-amino acid (aa) protein with a calculated molecular weight (Mr) of 42.9 kDa and an isoelectric point (pI) of 4.5 was characterized. The cDNAALP5 gene had an 18-amino acid signal peptide, one N-gylcosylation, one histidine active site, and one serine active site. The ALP5 gene with a Mr of 1351 bp contained two introns. One intron was of 54 bp, while the other was of 50 bp. Protein BLAST and phylogenetic tree analysis of the deduced amino sequences from the cDNAALP5 gene showed that the encoded protein had 100% homology to a protease enzyme (ALP2) of a sea strain of A. pullulans, suggesting that the protein ALP5 was an alkaline serine protease. Expression of ALP5 in Escherichia coli BL21 (DE3), followed by identification with Western-blotting, purification with Ni-NTA and analysis of enzymatic activity, yielded an homogeneous recombinant ALP5 which hydrolysed the substrate casein and inhibited the mycelial growth of the pathogens. At its optimal pH of 10.0 and reaction temperature of 50 °C, the recombinant protease exhibited the highest activity towards the substrate casein, though the highest stability was at lower temperatures and pH between 7.0 and 9.0. This study provided the direct evidence that extracellular proteases secreted by the antagonist A. pullulans PL5 played a role in the biocontrol activities against some postharvest pathogens of apple and peach. 相似文献
19.
A 15.0 kDa serine proteinase with collagenase activity from pyloric caeca of tuna, Thunnus thynnus, was purified in four steps; acetone precipitation, gel filtration chromatography on a Sephadex G‐100, ion‐exchange chromatography on a DEAE‐Sephadex α‐50 and gel filtration chromatography on a Sephadex G‐75 column. The purification and yield were 30.5‐fold and 0.023%, respectively, as compared with those in the starting crude extract. The optimum pH and temperature for the purified collagenolytic enzyme were around pH 7.5 and 55C, respectively. The purified proteinase was strongly inhibited by metal ions (Hg2+ and Zn2+) and serine proteinase inhibitors (PMSF, TLCK and soybean trypsin inhibitor) suggesting it is a serine protease. The Km and Vmax of the purified enzyme for collagen type I were approximately 3.82 mM and 851.5 U, respectively. 相似文献
20.
A dimeric serine protease Neriifolin S of molecular mass 94 kDa with milk clotting activity has been purified from the latex of Euphorbia neriifolia by anion exchange and size-exclusion chromatography. It hydrolyses peptidyl substrates l-Ala-pNA with highest affinity (Km of 0.195 mM) and physiological efficiency (Kcat/Km of 144.5 mM s). Enzyme belongs to the class of neutral proteases with pI value of 6.8, optimal proteolytic activity displayed at pH 9.5 and temperature 45 °C. Its proteolytic activity is strongly stimulated in the presence of Ca+2 ions and exclusively inhibited by serine protease inhibitors. Enzyme is fairly stable toward chemical denaturants, pH and temperature. The apparent Tm, was found to be 65 °C. Thermal inactivation follow first order kinetics with activation energy (Ea), activation enthalpy (ΔH∗), free energy change (ΔG∗) and entropy (ΔS∗) of 27.54 kJ mol−1, 24.89 kJ mol−1, −82.34 kJ mol−1 and 337.20 J mol−1 K−1. 相似文献