共查询到20条相似文献,搜索用时 15 毫秒
1.
Jian-Fen Zhang Yu-Guo Zheng Yin-Chu Shen 《Applied biochemistry and biotechnology》2010,162(4):966-974
A soluble 3-ketovalidoxylamine A C-N lyase from Stenotrophomonas maltrophilia was purified to 367.5-fold from the crude enzyme, with a yield of 16.4% by column chromatography on High S IEX, Methyl HIC, High Q IEX, and Sephadex G 100. The molecular mass of the enzyme was estimated to be 34 kDa by sodium dodecyl sulfate–polyacrylamide gel electrophoresis, and the enzyme was a neutral protein having an isoelectric point value at pH?7.0. The optimal pH of 3-ketovalidoxylamine A C-N lyase was around 7.0. The enzyme was stable within a pH range of 7.0–10.5. The optimal temperature was found to be near 40?°C, and the enzyme was sensitive to heat. The enzyme was completely inhibited by ethylenediaminetetraacetic acid, and it was reversed by Ca2+. The product, p-nitroaniline, inhibited the enzyme activity significantly at low concentration. The enzyme has C-N lyase activity and C-O lyase activity, and need 3-keto groups. The apparent K m value for p-nitrophenyl-3-ketovalidamine was 0.14 mM. 相似文献
2.
Zuming Li Zhihui Bai Baoguo Zhang Baojv Li Bo Jin Michael Zhang Francis Lin Hongxun Zhang 《Applied biochemistry and biotechnology》2012,167(8):2241-2256
Alkaline pectin lyase (PNL) shows potential as a biological control agent against several plant diseases. We isolated and characterized a new Bacillus clausii strain that can produce 4,180?U/g of PNL using sugar beet pulp as a carbon source and inducer. The PNL was purified to apparent homogeneity using ultrafiltration, ammonium sulfate fractionation, DEAE Sepharose Fast Flow, and Sephadex G-75 gel filtration. The purified PNL was found to be a monomeric protein with a molecular weight of 35?kDa, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). It demonstrated optimal activity with K m of 0.87?mg/ml at pH?10.0 and 60?°C. The enzyme is stable in the pH range of 8.0?C10.0 and temperature ??40?°C. Ca2+ was found to stimulate the enzymatic activity of the PNL by up to 410?%. Mass spectrometric results gave 38?% match coverage with pectate lyase from B. clausii KSM-K16 (gi|56961845). The PNL was found to elicit disease resistance in cucumber seedlings, suggesting that it may have applications in biocontrol and sustainable agriculture. 相似文献
3.
The purification and characterization of intracellular tyrosine phenol lyase from Citrobacter freundii has been carried out. The enzyme was purified 35-fold to homogeneity by ammonium sulphate precipitation and hydrophobic interaction chromatography. Its subunit molecular weight was found to be 52 kDa on sodium dodecyl sulphate polyacrylamide gel electrophoresis. The purified tyrosine phenol lyase showed maximum activity in borate buffer (0.05 M at pH 8.5) at 45 °C after 20 min of incubation. The K m and V max values of purified enzyme were found to be 0.446 mm and 0.342 mM/min/mg. This enzyme exhibits t 1/2 of 10, 52 and 130 min at 55, 45 and 35 °C, respectively. The N-terminal amino acid sequence was determined as MET-ASN-TYR-PRO-ALA-GLU-PRO-PHE-ARG-ILE-TRP-TRP-VAL-GLY. 相似文献
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5.
Nawel Boucherba Mohammed Gagaoua Estelle Copinet Azeddine Bettache Francis Duchiron Said Benallaoua 《Applied biochemistry and biotechnology》2014,172(5):2694-2705
Jonesia denitrificans BN-13 produces six xylanases: Xyl1, Xyl2, Xyl3, Xyl4, Xyl5, and Xyl6; the Xyl4 was purified and characterized after two consecutive purification steps using ultrafiltration and anion exchange chromatography. The xylanase-specific activity was found to be 77 unit (U)/mg. The molecular weight of the Xyl4 estimated using sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) revealed a monomeric isoenzyme of about 42 kDa. It showed an optimum pH value of 7.0 and a temperature of 50 °C. It was stable at 50 °C for 9.34 h. The enzyme showed to be activated by Mn+2, β-mercaptoethanol, and dithiothreitol (DTT) with a high affinity towards birchwood xylan (with a K m of 1 mg ml?1) and hydrolysis of oat-spelt xylan with a K m of 1.85 mg ml?1. The ability of binding to cellulose and/or xylan was also investigated. 相似文献
6.
Penicillium nalgiovense PNA9 produces an extracellular protease during fermentation with characteristics of growth-associated product. Enzyme purification involved ammonium sulfate precipitation, dialysis, and ultrafiltration, resulting in 12.1-fold increase of specific activity (19.5 U/mg). The protein was isolated through a series of BN-PAGE and native PAGE runs. ESI-MS analysis confirmed the molecular mass of 45.2 kDa. N-Terminal sequencing (MGFLKLLKGSLATLAVVNAGKLLTANDGDE) revealed 93 % similarity to a Penicillium chrysogenum protease, identified as major allergen. The protease exhibits simple Michaelis-Menten kinetics and K m (1.152 mg/ml), V max (0.827 mg/ml/min), and k cat (3.2?×?102) (1/s) values against azocasein show that it possesses high substrate affinity and catalytic efficiency. The protease is active within 10–45 °C, pH 4.0–10.0, and 0–3 M NaCl, while maximum activity was observed at 35 °C, pH 8.0, and 0.25 M NaCl. It is active against the muscle proteins actin and myosin and inactive against myoglobin. It is highly stable in the presence of non-ionic surfactants, hydrogen peroxide, BTNB, and EDTA. Activity was inhibited by SDS, Mn2+ and Zn2+, and by the serine protease inhibitor PMSF, indicating the serine protease nature of the enzyme. These properties make the novel protease a suitable candidate enzyme in meat ripening and other biotechnological applications. 相似文献
7.
C. E. Maepa J. Jayaramudu J. O. Okonkwo E. R. Sadiku J. Ramontja 《International Journal of Polymer Analysis and Characterization》2015,20(2):99-109
This article reports on the extraction and characterization of novel natural cellulose fibers obtained from the maize (tassel) plant. Cellulose was extracted from the agricultural residue (waste biomaterial) of maize tassel. The maize tassel fibers were obtained after treatment with NaOH and were carefully characterized while the chemical composition was determined. The chemical composition of the maize tassel fibers showed that the cellulose content increased from 41% to 56%, following alkali treatment. FT-IR spectroscopic analysis of maize tassel fibers confirmed that this chemical treatment also shows the way to partial elimination of hemicelluloses and lignin from the structure of the maize tassel fibers. X-ray diffraction results indicated that this process resulted in enhanced crystallinity of the maize tassel fibers. The thermal properties of the maize tassel fibers were studied by the TGA technique and were found to have improved significantly. The degradation temperature of the alkali-treated maize tassel fiber is higher than that of the untreated maize tassel fibers. This value convincingly showed the potential of maize tassel fibers for use in reinforced biocomposites and waste water treatment. 相似文献
8.
Nuttapon Songnaka Monthon Lertcanawanichakul Albert M. Hutapea Sucheewin Krobthong Yodying Yingchutrakul Apichart Atipairin 《Molecules (Basel, Switzerland)》2022,27(23)
Methicillin-resistant Staphylococcus aureus (MRSA) is listed as a high-priority pathogen because its infection is associated with a high mortality rate. It is urgent to search for new agents to treat such an infection. Our previous study isolated a soil bacterium (Brevibacillus sp. SPR-20), showing the highest antimicrobial activity against S. aureus TISTR 517 and MRSA strains. The present study aimed to purify and characterize anti-MRSA substances produced by SPR-20. The result showed that five active substances (P1–P5) were found, and they were identified by LC-MS/MS that provided the peptide sequences of 14–15 residues. Circular dichroism showed that all peptides contained β-strand and disordered conformations as the major secondary structures. Only P1–P4 adopted more α-helix conformations when incubated with 50 mM SDS. These anti-MRSA peptides could inhibit S. aureus and MRSA in concentrations of 2–32 μg/mL. P1 (NH2-VVVNVLVKVLPPPVV-COOH) had the highest activity and was identified as a novel antimicrobial peptide (AMP). The stability study revealed that P1 was stable in response to temperature, proteolytic enzymes, surfactant, and pH. The electron micrograph showed that P1 induced bacterial membrane damage when treated at 1× MIC in the first hour of incubation. The killing kinetics of P1 was dependent on concentration and time. Mechanisms of P1 on tested pathogens involved membrane permeability, leakage of genetic material, and cell lysis. The P1 peptide at a concentration up to 32 μg/mL showed hemolysis of less than 10%, supporting its safety for human erythrocytes. This study provides promising anti-MRSA peptides that might be developed for effective antibiotics in the post-antibiotic era. 相似文献
9.
Bruno C. van der Aa Marcel Asther Yves F. Dufrêne 《Colloids and surfaces. B, Biointerfaces》2002,24(3-4):277-284
Spores of the filamentous fungus Aspergillus oryzae have a great biotechnological potential for the production of highly active proteins. To date, little is known about the molecular mechanisms of spore aggregation, a phenomenon observed during germination in liquid medium. Here, atomic force microscopy (AFM) imaging and force measurements were used to characterize, under aqueous conditions, the surface morphology and macromolecular interactions of A. oryzae spores in relation to their aggregation behavior. Dormant spores were covered with a discontinuous layer of about 35 nm thickness, as revealed by height images. High-resolution deflection images showed that this layer consisted of rodlets, 10±1 nm in diameter, that were assembled in parallel to form fascicles interlaced with different orientations. The germinating spore surface was much rougher and showed streaks oriented in the scanning direction, indicating that the probe was interacting with soft material. Retraction force curves were strikingly different depending on the spore physiological state: while dormant spores exhibited non-adhesive properties, germinating spores showed single or multiple attractive forces of 400±100 pN magnitude, along with characteristic elongation forces and rupture lengths ranging from 20 to 500 nm. These elongation forces are attributed to the stretching of long, flexible cell surface macromolecules and suggested to play a role in the aggregation process by promoting bridging interactions. 相似文献
10.
兽疫链球菌变异株产生的透明质酸的纯化及表征 总被引:6,自引:0,他引:6
用N-甲基-N′-硝基-N-亚硝基胍(NTG)对兽疫链球菌进行诱变,获得高产菌株.经过对该菌株的发酵培养,将产生的多糖经Savage法、季铵复合物沉淀法、DEAE-纤维素(DE52)离子交换层析法及SephadexG-75凝胶过滤法分离纯化.纯化的多糖结构经化学组成分析、核磁共振波谱、红外光谱及圆二色谱鉴定,证明了诱变得到的高产菌株(StreptococcuszooepidemicusJ18)再经发酵,得到的多糖为透明质酸.通过刚果红实验证明了透明质酸的构象为单股螺旋结构,其平均分子量约为1.16×106. 相似文献
11.
Bao Zhang Chunjuan Dong Qingmao Shang Yuan Cong Weijia Kong Pinglan Li 《Applied biochemistry and biotechnology》2013,171(8):2262-2272
Bacillus amyloliquefaciens K103 isolated from a lemon sample was used as a biocontrol agent to suppress Rhizoctonia solani Kühn and other fungal plant pathogens. Two antifungal compounds were purified from the culture broth using acid precipitation, gel permeation chromatography, and reversed-phase high-performance liquid chromatography. Matrix-assisted laser desorption ionization-time of flight mass spectrometry analysis indicated that the antifungal compounds were two isomers similar to bacillomycin L. One of the predominant active fractions was subjected to quadrupole time-of-flight mass spectrometry and amino acid analysis to determine its structural characteristics, revealing that the antifungal compound with a molecular mass of 1,034.5464 was identical to bacillomycin L. This is the second report of lemon microflora producing bacillomycin L or any antifungal compound, suppressing the growth of R. solani Kühn. Meanwhile, the study provided insights into the enormous potential of food microbial resources and bacillomycin L antibiotics in biological control and sustainable agriculture. 相似文献
12.
Melt spinning of poly-L, DL-lactide 70/30 has been studied. Fiber having diameter lower than 120 micron exhibited tensile modulus and strength in the range of 3–4 GPa and 130–180 MPa, respectively. Maximum attainable modulus and strength of 4.7 GPa and 205 MPa were predicted, according to a proposed equation in dependence on the draw ratio. In vitro degradation performed in PBS solution at 37 °C, showed that after 4 weeks fibers maintained adequate properties for tissue engineering applications. 相似文献
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Asiya Nazir Rohit Soni H. S. Saini Amarjeet Kaur B. S. Chadha 《Applied biochemistry and biotechnology》2010,162(2):538-547
This study reports differential expression of endoglucanase (EG) and β-glucosidase (βG) isoforms of Aspergillus terreus. Expression of multiple isoforms was observed, in presence of different carbon sources and culture conditions, by activity
staining of poly acrylamide gel electrophoresis gels. Maximal expression of four EG isoforms was observed in presence of rice
straw (28 U/g DW substrate) and corn cobs (1.147 U/ml) under solid substrate and shake flask culture, respectively. Furthermore,
the sequential induction of EG isoforms was found to be associated with the presence of distinct metabolites (monosaccharides/oligosaccharides)
i.e., xylose (X), G1, G3 and G4 as well as putative positional isomers (G1/G2, G2/G3) in the culture extracts sampled at different time intervals, indicating specific role of these metabolites in the sequential
expression of multiple EGs. Addition of fructose and cellobiose to corn cobs containing medium during shake flask culture
resulted in up-regulation of EG activity, whereas addition of mannitol, ethanol and glycerol selectively repressed the expression
of three EG isoforms (Ia, Ic and Id). The observed regulation profile of βG isoforms was distinct when compared to EG isoforms,
and addition of glucose, fructose, sucrose, cellobiose, mannitol and glycerol resulted in down-regulation of one or more of
the four βG isoforms. 相似文献
15.
Andre Ricardo de Lima Damásio Tony Márcio da Silva Alexandre Maller Jo?o Atílio Jorge Hector Francisco Terenzi Maria de Lourdes Teixeira de Moraes Polizeli 《Applied biochemistry and biotechnology》2010,160(5):1496-1507
An extracellular polygalacturonase (PG) produced from Paecilomyces variotii was purified to homogeneity through two chromatography steps using DEAE-Fractogel and Sephadex G-100. The molecular weight
of P. variotii PG was 77,300 Da by gel filtration and SDS-PAGE. PG had isoelectric point of 4.37 and optimum pH 4.0. PG was very stable
from pH 3.0 to 6.0. The extent of hydrolysis of different pectins by the purified enzyme was decreased with an increase in
the degree of esterification. PG had no activity toward non-pectic polysaccharides. The apparent K
m and V
max values for hydrolyzing sodium polypectate were 1.84 mg/mL and 432 μmol/min/mg, respectively. PG was found to have temperature
optimum at 65 °C and was totally stable at 45 °C for 90 min. Half-life at 55 °C was 50.6 min. Almost all the examined metal
cations showed partial inhibitory effects under enzymatic activity, except for Na+1, K+1, and Co+2 (1 mM) and Cu+2 (1 and 10 mM). 相似文献
16.
Sara Tombelli Marco Mascini Barbara Scherm Gianni Battacone Quirico Migheli 《Monatshefte für Chemie / Chemical Monthly》2009,67(2):901-907
Abstract
In this work, we report on the development of a DNA-based piezoelectric biosensor specific for the detection of an amplicon of the aflD gene of Aspergillus flavus and A. parasiticus. Expression of this gene is consistently correlated with a strain’s ability to produce aflatoxins that are considered very potent liver carcinogens in various animal species and humans. The DNA biosensor has been characterized with synthetic oligonucleotides and amplicons. Moreover, it has been applied to the analysis of real samples consisting of amplicons of DNA extracted from flours and feed contaminated with A. flavus and A. parasiticus. 相似文献17.
LIU Yang LI Quan-shun ZHU Hong-liang MENG Zhao-li XIANG Hong-yu XIE Qiu-hong 《高等学校化学研究》2013,29(5):917-923
Two thermostable glucoamylases were produced from Aspergillus niger B-30 by submerged fermentation. The two glucoamylases GAM-1 and GAM-2 were purified by ammonium sulfate precipitation, diethylaminoethylcellulose fast flow(DEAE FF) and Superdex G-75 gel filtration columns. The molecular weights of GAM-1 and GAM-2 were determined as 9.72×104 and 7.83×104 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), while the molecular weights of GAM-1 and GAM-2 were determined to be 8.05×104 and 7.04×104 by matrix assisted laser desorption ionizationtime-of-flight(MALDI-TOF) mass spectrometry, respectively. Both the enzymes were glycosylated, with 10.4% and 11.4% carbohydrate content, respectively. The optimal pH and temperature were 4.0―4.6 and 70 ℃ for both. The two glucoamylases were maintained 100% relative activity after incubation at 60 ℃ for 120 min. After the hydrolysis of starch for 120 min, glucose was the only product, confirming that the two enzymes were of high efficiency towards starch. The GAM-2 exhibited higher catalytic activity towards oligosaccharides such as maltose than GAM-1, and the kinetic analysis shows that the affinity of GAM-2 to starch was lower than that of GAM-1. The high thermostability and effectiveness make the two glucoamylases potentially attractive for biotechnological application. 相似文献
18.
Sree Renjini Isac Gopinath Balakrish Nair Durg Vijai Singh 《Applied biochemistry and biotechnology》2012,167(4):809-823
Vibrio cholerae O54 TV113 isolated from a diarrheal patient produces an extracellular cytotoxin that caused alteration in the morphology of Chinese hamster ovary cells manifested as cell shrinkage with intact cell boundaries and finally causing cell death. Syncase medium supplemented with lincomycin (50???g/ml), pH 7.2, and 18?h incubation with shaking at 37?°C supported optimal cytotoxin production. We isolated and purified this cytotoxin to homogeneity by ultrafiltration, 40?C80?% ammonium sulfate precipitation, gradient?Canion exchange chromatography, stepwise-anion exchange chromatography, and size exclusion chromatography increasing the specific activity by 866-fold. The cytotoxin is heat-labile, sensitive to protease and papain, and has a molecular weight of 64?kDa determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and enterotoxic activity in rabbit ileal loop assay. Both cytotoxic and enterotoxic activity could be inhibited or neutralized by antiserum raised against purified cytotoxin but not by preimmune serum. Immunodiffusion test between purified cytotoxin and its antiserum gave a single well-defined precipitin band showing reaction of complete identity and a well-defined single band in an immunoblot assay. This study thus indicate that the cytotoxin expressed by strain TV113 has both cytotoxic and enterotoxic activity and appears to contribute in pathogenesis of non-O1, non-O139 strains. 相似文献
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