共查询到20条相似文献,搜索用时 312 毫秒
1.
M. López-Sánchez M. J. Ayora-Cañada A. Molina-Díaz M. Siam W. Huber G. Quintás S. Armenta B. Lendl 《Analytical and bioanalytical chemistry》2009,394(8):2137-2144
A mid-infrared enzymatic assay for label-free monitoring of the enzymatic reaction of fructose-1,6-bisphosphatase with fructose
1,6-bisphosphate has been proposed. The whole procedure was done in an automated way operating in the stopped flow mode by
incorporating a temperature-controlled flow cell in a sequential injection manifold. Fourier transform infrared difference
spectra were evaluated for kinetic parameters, like the Michaelis–Menten constant (K
M) of the enzyme and V
max of the reaction. The obtained K
M of the reaction was 14 ± 3 g L−1 (41 μM). Furthermore, inhibition by adenosine 5′-monophosphate (AMP) was evaluated, and the K
MApp value was determined to be 12 ± 2 g L−1 (35 μM) for 7.5 and 15 μM AMP, respectively, with V
max decreasing from 0.1 ± 0.03 to 0.05 ± 0.01 g L−1 min−1. Therefore, AMP exerted a non-competitive inhibition. 相似文献
2.
Wanli Liu Pengjun Shi Qiang Chen Peilong Yang Guozeng Wang Yaru Wang Huiying Luo Bin Yao 《Applied biochemistry and biotechnology》2010,162(1):1-12
A xylanase-encoding gene, xyn11F63, was isolated from Penicillium sp. F63 CGMCC1669 using degenerated polymerase chain reaction (PCR) and thermal asymmetric interlaced (TAIL)-PCR techniques.
The full-length chromosomal gene consists of 724 bp, including a 73-bp intron, and encodes a 217 amino acid polypeptide. The
deduced amino acid sequence of xyn11F63 shows the highest identity of 70% to the xylanase from Penicillium sp. strain 40, which belongs to glycosyl hydrolases family 11. The gene was overexpressed in Pichia pastoris, and its activity in the culture medium reached 516 U ml−1. After purification to electrophoretic homogeneity, the enzyme showed maximal activity at pH 4.5 and 40°C, was stable at
acidic buffers of pH 4.5–9.0, and was resistant to proteases (proteinase K, trypsin, subtilisin A, and α-chymotrypsin). The
specific activity, K
m, and V
max for oat spelt xylan substrate was 7,988 U mg−1, 22.2 mg ml−1, and 15,105.7 μmol min−1 mg−1, respectively. These properties make XYN11F63 a potential economical candidate for use in feed and food industrial applications. 相似文献
3.
Wei Zhao Aisheng Xiong Xiaoyan Fu Feng Gao Yongsheng Tian Rihe Peng 《Applied biochemistry and biotechnology》2010,162(8):2157-2165
To obtain a high level expression of phytase with favorable characteristics, a codon-optimized phytase gene from Citrobacter freundii was synthesized and transferred into Pichia pastoris. Small-scale expression experiments and activity assays were used to screen positive colonies. After purified by Ni2+–NTA agarose affinity column, the characterizations of the recombinant phytase were determined. The recombinant phytase (r-phyC)
had two distinct pH optima at 2.5 and 4.5 and an optimal temperature at 50 °C. It retained more than 80% activity after being
incubated under various buffer (pH 1.5–8.0) at 37 °C for 1 h. The specific activity, Km, and Vmax values of r-phyC for sodium phytate were 2,072 ± 18 U mg−1, 0.52 ± 0.04 mM, and 2,380 ± 84 U mg−1 min−1, respectively. The enzyme activity was significantly improved by 1 mM of K+, Ca2+, and Mg2+. These characteristics contribute to its potential application in feed industry. 相似文献
4.
Wanwisa Moon-ai Ploypat Niyomploy Ruethairat Boonsombat Polkit Sangvanich Aphichart Karnchanatat 《Applied biochemistry and biotechnology》2012,166(8):2138-2155
Superoxide dismutase (SOD, EC 1.15.1.1) is a metalloenzyme or antioxidant enzyme that catalyzes the disproportionation of
the harmful superoxide anionic radical to hydrogen peroxide and molecular oxygen. Due to its antioxidative effects, SOD has
long been applied in medicinal treatment, cosmetic, and other chemical industries. Fifteen Zingiberaceae plants were tested
for SOD activity in their rhizome extracts. The crude homogenate and ammonium sulfate cut fraction of Curcuma aeruginosa were found to contain a significant level of SOD activity. The SOD enzyme was enriched 16.7-fold by sequential ammonium sulfate
precipitation, diethylaminoethyl cellulose ion exchange, and Superdex 75 gel filtration column chromatography. An overall
SOD yield of 2.51 % with a specific activity of 812.20 U/mg was obtained. The enriched SOD had an apparent MW of 31.5 kDa,
as judged by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and a pH and temperature optima of 4.0 and 50 °C.
With nitroblue tetrazolium and riboflavin as substrates, the K
m values were 57.31 ± 0.012 and 1.51 ± 0.014 M, respectively, with corresponding V
max values of 333.7 ± 0.034 and 254.1 ± 0.022 μmol min−1 mg protein−1. This SOD likely belongs to the Fe- or Mn-SOD category due to the fact that it was insensitive to potassium cyanide or hydrogen
peroxide inhibition, but was potentially weakly stimulated by hydrogen peroxide, and stimulated by Mn2+and Fe2+ ions. Moreover, this purified SOD also exhibited inhibitory effects on lipopolysaccharide-induced nitric oxide production
in cultured mouse macrophage cell RAW 264.7 in a dose-dependent manner (IC50 = 14.36 ± 0.15 μg protein/ml). 相似文献
5.
Javed MR Rashid MH Nadeem H Riaz M Perveen R 《Applied biochemistry and biotechnology》2009,157(3):483-497
Monomeric extracellular endoglucanase (25 kDa) of transgenic koji (Aspergillus oryzae cmc-1) produced under submerged growth condition (7.5 U mg−1 protein) was purified to homogeneity level by ammonium sulfate precipitation and various column chromatography on fast protein
liquid chromatography system. Activation energy for carboxymethylcellulose (CMC) hydrolysis was 3.32 kJ mol−1 at optimum temperature (55 °C), and its temperature quotient (Q
10) was 1.0. The enzyme was stable over a pH range of 4.1–5.3 and gave maximum activity at pH 4.4. V
max for CMC hydrolysis was 854 U mg−1 protein and K
m was 20 mg CMC ml−1. The turnover (k
cat) was 356 s−1. The pK
a1 and pK
a2 of ionisable groups of active site controlling V
max were 3.9 and 6.25, respectively. Thermodynamic parameters for CMC hydrolysis were as follows: ΔH* = 0.59 kJ mol−1, ΔG* = 64.57 kJ mol−1 and ΔS* = −195.05 J mol−1 K−1, respectively. Activation energy for irreversible inactivation ‘E
a(d)’ of the endoglucanase was 378 kJ mol−1, whereas enthalpy (ΔH*), Gibbs free energy (ΔG*) and entropy (ΔS*) of activation at 44 °C were 375.36 kJ mol−1, 111.36 kJ mol−1 and 833.06 J mol−1 K−1, respectively. 相似文献
6.
Inna S. Alpeeva Aiste Vilkanauskyte Bertrand Ngounou Elisabeth Csöregi Ivan Yu. Sakharov Mykhailo Gonchar Wolfgang Schuhmann 《Mikrochimica acta》2005,152(1-2):21-27
We report on the development of a bi-layer bi-enzyme biosensor architecture using different peroxidases and alcohol oxidase
from Hansenula polymorpha C-105 as biological recognition elements. The sensor architecture comprises a first layer containing either horseradish peroxidase
or royal palm tree peroxidase crosslinked with an Osmium complex-modified redox hydrogel. On top, a second layer was formed
by electrochemically induced precipitation of a cathodic electrodeposition paint simultaneously entrapping alcohol oxidase
isolated from a genetically modified strain of Hansenula polymorpha C-105. The sensor architecture was optimized with respect to effective electron transfer and stability of the enzyme. The
main characteristics of the biosensors are an apparent maximal current Imaxapp of 572–940 nA, an apparent Michaelis constant KMapp of 9.5 mM, a sensitivity of 60–98 nA mM−1 and an improved operational stability represented by a deactivation constant of 1.5–2.0 × 10−4min−1. 相似文献
7.
Silvia Castrignanò Sheila J. Sadeghi Gianfranco Gilardi 《Analytical and bioanalytical chemistry》2010,398(3):1403-1409
Human flavin-containing monooxygenases are the second most important class of drug-metabolizing enzymes after cytochromes
P450. Here we report a simple but functional and stable enzyme-electrode system based on a glassy carbon (GC) electrode with
human flavin-containing monooxygenase isoform 3 (hFMO3) entrapped in a gel cross-linked with bovine serum albumin (BSA) by
glutaraldehyde. The enzymatic electrochemical responsiveness is characterised by using well-known substrates: trimethylamine
(TMA), ammonia (NH3), triethylamine (TEA), and benzydamine (BZD). The apparent Michaelis–Menten constant (K′M) and apparent maximum current (I′max) are calculated by fitting the current signal to the Michaelis–Menten equation for each substrate. The enzyme-electrode has
good characteristics: the calculated sensitivity was 40.9 ± 0.5 mA mol−1 L cm−2 for TMA, 43.3 ± 0.1 mA mol−1 L cm−2 for NH3, 45.2 ± 2.2 mA mol−1 L cm−2 for TEA, and 39.3 ± 0.6 mA mol−1 L cm−2 for BZD. The stability was constant for 3 days and the inter-electrode reproducibility was 12.5%. This is a novel electrochemical
tool that can be used to investigate new potential drugs against the catalytic activity of hFMO3. 相似文献
8.
Pengjun Shi Ning Li Peilong Yang Yaru Wang Huiying Luo Yingguo Bai Bin Yao 《Applied biochemistry and biotechnology》2010,162(3):707-718
An α-l-arabinofuranosidase gene, abf51S9, was cloned from Streptomyces sp. S9 and successfully expressed in Escherichia coli BL21 (DE3). The full-length gene consisted of 1,506 bp and encoded 501 amino acids with a calculated mass of 55.2 kDa. The
deduced amino acid sequence was highly homologous with the α-l-arabinofuranosidases belonging to family 51 of the glycoside hydrolases. The recombinant protein was purified to electrophoretic
homogeneity by Ni-NTA affinity chromatography and subsequently characterized. The optimal pH and temperature for the recombinant
enzyme were 6.0 and 60∼65 °C, respectively. The enzyme showed a broad pH range of stability, retaining over 75% of the maximum
activity at pH 5.0 to 11.0. The specific activity, K
m, and V
max with p-nitrophenyl-α-l-arabinofuranoside as substrate were 60.0 U mg−1, 1.45 mM, and 221 μmol min−1 mg−1, respectively. Abf51S9 showed a mild but significant synergistic effect in combination with xylanase on the degradation of
oat-spelt xylan and soluble wheat arabinoxylan substrates with a 1.19- and 1.21-fold increase in the amount of reducing sugar
released, respectively. These favorable properties make Abf51S9 a good candidate in various industrial applications. 相似文献
9.
Qiong Liu Peilong Yang Huiying Luo Pengjun Shi Huoqing Huang Kun Meng Bin Yao 《Applied biochemistry and biotechnology》2012,166(6):1442-1453
An endo-β-1,4-mannanase encoding gene, man5, was cloned from Bispora antennata CBS 126.38, which was isolated from a beech stump. The cDNA of man5 consists of 1,299 base pairs and encodes a 432-amino-acid protein with a theoretical molecular mass of 46.6 kDa. Deduced
MAN5 exhibited the highest amino acid sequence identity of 58% to a β-mannanase of glycoside hydrolase family 5 from Aspergillus aculeatus. Recombinant MAN5 was expressed in Pichia pastoris and purified to electrophoretic homogeneity. The specific activity of the final preparation towards locust bean gum was 289
U mg−1. MAN5 showed optimal activity at pH 6.0 and 70 °C and had good adaptation and stability over a broad range of pH values.
The enzyme showed more than 60% of peak activity at pH 3.0–8.0 and retained more than 80% of activity after incubation at
37 °C for 1 h in both acid and alkaline conditions (pH 4.0–11.0). The K
m and V
max values were 1.33 mg ml−1 and 444 μmol min−1 mg−1 and 1.17 mg ml−1 and 196 μmol min−1 mg−1 for locust bean gum and konjac flour, respectively. Of all tested metal ions and chemical reagents, Co2+, Ni2+, and β-mercaptoethanol enhanced the enzyme activity at 1 mM, whereas other chemicals had no effect on or partially inhibited
the enzyme activity. MAN5 was highly resistant to acidic and neutral proteases (trypsin, α-chymotrypsin, collagenase, subtilisin
A, and proteinase K). By virtue of the favorable properties of MAN5, it is possible to apply this enzyme in the paper and
food industries. 相似文献
10.
Vivekanand V Dwivedi P Pareek N Singh RP 《Applied biochemistry and biotechnology》2011,165(1):204-220
In solid-state fermentation, among various solid supports evaluated, banana peel was found to be an ideal support and resulted
into higher levels of laccase (6281.4 ± 63.60 U l−1) along with notable levels of manganese peroxidase production (1339.0 ± 131.23 U l−1) by Aspergillus fumigatus VkJ2.4.5. Maximum levels of laccase was achieved under derived conditions consisting of 80% of moisture level, 6 days of
incubation period, 6% inoculum level, and an aeration level of 2.5 l min−1. A column-tray bioreactor was designed to scale up and economize the enzyme production in three successive cycles of fermentation
using the same fungal biomass. Thermal and pH stability profiles revealed that enzyme was stable up to 50°C and at varying
pH range from 5–9 for up to 2 h. The apparent molecular weight of laccase was found to be 34 ± 1 kDa. MALDI-TOF/TOF analysis
of the protein showed significant homology with maximum identity of 67% to other laccases reported in database. 相似文献
11.
Maalej I Belhaj I Masmoudi NF Belghith H 《Applied biochemistry and biotechnology》2009,158(1):200-212
A thermostable xylanase from a newly isolated thermophilic fungus Talaromyces thermophilus was purified and characterized. The enzyme was purified to homogeneity by ammonium sulfate precipitation, diethylaminoethyl
cellulose anion exchange chromatography, P-100 gel filtration, and Mono Q chromatography with a 23-fold increase in specific
activity and 17.5% recovery. The molecular weight of the xylanase was estimated to be 25kDa by sodium dodecyl sulfate–polyacrylamide
gel electrophoresis and gel filtration. The enzyme was highly active over a wide range of pH from 4.0 to 10.0. The relative
activities at pH5.0, 9.0, and 10.0 were about 80%, 85.0%, and 60% of that at pH7.5, respectively. The optimum temperature
of the purified enzyme was 75°C. The enzyme showed high thermal stability at 50°C (7days) and the half-life of the xylanase
at 100°C was 60min. The enzyme was free from cellulase activity. K
m and V
max values at 50°C of the purified enzyme for birchwood xylan were 22.51mg/ml and 1.235μmol min−1 mg−1, respectively. The enzyme was activated by Ag+, Co2+, and Cu2+; on the other hand, Hg2+, Ba2+, and Mn2+ inhibited the enzyme. The present study is among the first works to examine and describe a secreted, cellulase-free, and
highly thermostable xylanase from the T. thermophilus fungus whose application as a pre-bleaching aid is of apparent importance for pulp and paper industries. 相似文献
12.
S. Subramaniyan 《Applied biochemistry and biotechnology》2012,166(7):1831-1842
Low molecular weight endo-xylanase from Bacillus pumilus SSP-34 was purified to homogeneity using ion exchange and size exclusion chromatographies. Xylanases were isolated by novel
purification protocol which includes the use of anion exchange matrix such as DEAE Sepharose CL 6B with less affinity towards
enzyme protein. The purified B. pumilus SSP-34 have a molecular weight of 20 kDa, with optimum pH and temperature at 6.0 and 50 °C, respectively. The enzyme was
stable at 50 °C for 30 min. It showed remarkable stability at pH values ranging from 4.5 to 9 when the reaction was carried
out at 50 °C. K
m and V
max values, determined with oats spelts xylan were 6.5 mg ml−1 and 1,233 μmol min−1 mg−1 protein, respectively, and the specific activity was 1,723 U mg−1 相似文献
13.
A fast and sensitive liquid chromatography–mass spectrometry method was developed for the determination of ursolic acid (UA)
in rat plasma and tissues. Glycyrrhetinic acid was used as the internal standard (IS). Chromatographic separation was performed
on a 3.5 μm Zorbax SB-C18 column (30 mm × 2.1 mm) with a mobile phase consisting of methanol and aqueous 10 mM ammonium acetate
using gradient elution. Quantification was performed by selected ion monitoring with (m/z)− 455 for UA and (m/z)− 469 for the IS. The method was validated in the concentration range of 2.5 − 1470 ng mL−1 for plasma samples and 20 − 11760 ng g−1 for tissue homogenates. The intra- and inter-day assay of precision in plasma and tissues ranged from 1.6% to 7.1% and 3.7%
to 9.0%, respectively, and the intra- and inter-day assay accuracy was 84.2 − 106.9% and 82.1 − 108.1%, respectively. Recoveries
in plasma and tissues ranged from 83.2% to 106.2%. The limits of detections were 0.5 ng mL−1 or 4.0 ng g−1. The recoveries for all samples were >90%, except for liver, which indicated that ursolic acid may metabolize in liver. The
main pharmacokinetic parameters obtained were T
max = 0.42 ± 0.11 h, C
max = 1.10 ± 0.31 μg mL−1, AUC = 1.45 ± 0.21 μg h mL−1 and K
a = 5.64 ± 1.89 h−1. The concentrations of UA in rat lung, spleen, liver, heart, and cerebellum were studied for the first time. This method
is validated and could be applicable to the investigation of the pharmacokinetics and tissue distribution of UA in rats. 相似文献
14.
Mário A. P. Nunes Hélder Vila-Real Pedro C. B. Fernandes Maria H. L. Ribeiro 《Applied biochemistry and biotechnology》2010,160(7):2129-2147
A synthetic polymer, polyvinyl alcohol (PVA), a cheap and nontoxic synthetic polymer to organism, has been ascribed for biocatalyst
immobilization. In this work PVA–alginate beads were developed with thermal, mechanical, and chemical stability to high temperatures
(<80 °C). The combination of alginate and bead treatment with sodium sulfate not only prevented agglomeration but produced
beads of high gel strength and conferred enzyme protection from inactivation by boric acid. Naringinase from Penicillium decumbens was immobilized in PVA (10%)–alginate beads with three different sizes (1–3 mm), at three different alginate concentrations
(0.2–1.0%), and these features were investigated in terms of swelling ratio within the beads, enzyme activity, and immobilization
yield during hydrolysis of naringin. The pH and temperature optimum were 4.0 and 70 °C for the PVA–alginate-immobilized naringinase.
The highest naringinase activity yield in PVA (10%)–alginate (1%) beads of 2 mm was 80%, at pH 4.0 and 70 °C. The Michaelis
constant (K
Mapp) and the maximum reaction velocity (V
maxapp) were evaluated for both free (K
Mapp = 0.233 mM; V
maxapp = 0.13 mM min−1) and immobilized naringinase (K
Mapp = 0.349 mM; V
maxapp = 0.08 mM min−1). The residual activity of the immobilized enzyme was followed in eight consecutive batch runs with a retention activity
of 70%. After 6 weeks, upon storage in acetate buffer pH 4 at 4 °C, the immobilized biocatalyst retained 90% of the initial
activity. These promising results are illustrative of the potential of this immobilization strategy for the system evaluated
and suggest that its application may be effectively performed for the entrapment of other biocatalysts. 相似文献
15.
Yi Wang Jian Zhao Jian-He Xu Li-Qiang Fan Su-Xia Li Li-Li Zhao Xiao-Bo Mao 《Applied biochemistry and biotechnology》2010,162(8):2387-2399
A lipase gene from Serratia marcescens ECU1010 was cloned into expression vector pET28a, sequenced, and overexpressed as an N terminus His-tag fusion protein in
Escherichia coli. Through the optimization of culture conditions in shake flask, the lipase activity was improved up to 1.09 × 105 U/l, which is a great improvement compared to our previous reports. It was purified to homogeneity by Ni-NTA affinity chromatography
with an overall yield of 59.4% and a purification factor of 2.4-fold. This recombinant lipase displayed excellent stability
below 30 °C and within the pH range of 5.0−6.8, giving temperature and pH optima at 40 °C and pH 9.0, respectively. The lipase
activity was found to increase in the presence of metal ions such as Ca2+, Cu2+, and some nonionic surfactants such as PEG series. In addition, among p-nitrophenyl esters of fatty acids with varied chain length, the recombinant lipase showed the maximum activity on p-nitrophenyl laurate (C12). Using racemic trans-3-(4′-methoxy-phenyl)-glycidyl methyl ester [(±)-MPGM] as substrate, which is a key chiral synthon for production of diltiazem,
a 50% conversion yield was achieved after 4 h in toluene–water (100 mM KPB phosphate buffer, pH 7.5) biphasic system (5:5 ml)
at 30 °C under shaking condition (160 rpm), affording (−)-MPGM in nearly 100% ee. The K
m and V
max values of the lipase for (±)-MPGM were 222 mM and 1.24 mmol min−1 mg−1, respectively. The above-mentioned features make the highly enantioselective lipase from Serratia marcescens ECU1010 a robust biocatalyst for practical use in large-scale production of diltiazem intermediate. 相似文献
16.
Radix Scrophulariae (Xuanshen) is one of the famous Chinese herbal medicines widely used to treat rheumatism, tussis, pharyngalgia, arthritis,
constipation, and conjunctival congestion. Harpagoside and cinnamic acid are the main bioactive components of Xuanshen. The
purpose of this study was to develop an HPLC–UV method for simultaneous determination of harpagoside and cinnamic acid in
rat plasma and investigate pharmacokinetic parameters of harpagoside and cinnamic acid after oral administration of Xuanshen
extract (760 mg kg−1). After addition of syringin as internal standard, the analytes were isolated from plasma by liquid–liquid extraction. Separation
was achieved on a Kromasil C18 column, and detection was by UV absorption at 272 nm. The described assay was validated in terms of linearity, accuracy,
precision, recovery, and limit of quantification according to the FDA validation guidelines. Calibration curves for both analytes
were linear with the coefficient of variation (r) for both was greater than 0.999. Accuracy for harpagoside and cinnamic acid ranged from 100.7–103.5% and 96.9–102.9%, respectively,
and precision for both analytes were less than 8.5%. The main pharmacokinetic parameters found for harpagoside and cinnamic
acid after oral infusion of Xuanshen extract were as follows: C
max 1488.7 ± 205.9 and 556.8 ± 94.2 ng mL−1, T
max 2.09 ± 0.31 and (1.48 ± 0.14 h, AUC0–24 10336.4 ± 1426.8 and 3653.1 ± 456.4 ng h mL−1, 11276.8 ± 1321.4 and 3704.5 ± 398.8 ng h mL−1, and t
1/2 4.9 ± 1.3 and 2.5 ± 0.9 h, respectively. These results indicated that the proposed method is simple, selective, and feasible
for pharmacokinetic study of Radix Scrophulariae extract in rats.
Figure Radix Scrophulariae 相似文献
17.
Dengeti Shrinivas Gunashekaran Savitha Kumar Raviranjan Gajanan Ramchandra Naik 《Applied biochemistry and biotechnology》2010,162(7):2049-2057
A highly thermostable alkaline xylanase was purified to homogeneity from culture supernatant of Bacillus sp. JB 99 using DEAE-Sepharose and Sephadex G-100 gel filtration with 25.7-fold increase in activity and 43.5% recovery.
The molecular weight of the purified xylanase was found to be 20 kDA by SDS-PAGE and zymogram analysis. The enzyme was optimally
active at 70 °C, pH 8.0 and stable over pH range of 6.0–10.0.The relative activity at 9.0 and 10.0 were 90% and 85% of that
of pH 8.0, respectively. The enzyme showed high thermal stability at 60 °C with 95% of its activity after 5 h. The K
m and V
max of enzyme for oat spelt xylan were 4.8 mg/ml and 218.6 μM min−1 mg−1, respectively. Analysis of N-terminal amino acid sequence revealed that the xylanase belongs to glycosyl hydrolase family
11 from thermoalkalophilic Bacillus sp. with basic pI. Substrate specificity showed a high activity on xylan-containing substrate and cellulase-free nature. The hydrolyzed product
pattern of oat spelt xylan on thin-layer chromatography suggested xylanase as an endoxylanase. Due to these properties, xylanase
from Bacillus sp. JB 99 was found to be highly compatible for paper and pulp industry. 相似文献
18.
Rodrigues TH Rocha MV de Macedo GR Gonçalves LR 《Applied biochemistry and biotechnology》2011,164(6):929-943
In this work, the potential of microwave-assisted alkali pretreatment in order to improve the rupture of the recalcitrant
structures of the cashew able bagasse (CAB), lignocellulosic by-product in Brazil with no commercial value, is obtained from
cashew apple process to juice production, was studied. First, biomass composition of CAB was determined, and the percentage
of glucan and lignin was 20.54 ± 0.70% and 33.80 ± 1.30%, respectively. CAB content in terms of cellulose, hemicelluloses,
and lignin, 19.21 ± 0.35%, 12.05 ± 0.37%, and 38.11 ± 0.08%, respectively, was also determined. Results showed that, after
enzymatic hydrolysis, alkali concentration exerted influence on glucose formation, after pretreatment with 0.2 and 1.0 mo L−1 of NaOH (372 ± 12 and 355 ± 37 mg gglucan−1) when 2% (w/v) of cashew apple bagasse pretreated by microwave-assisted alkali pretreatment (CAB-M) was used. On the other hand, pretreatment
time (15–30 min) and microwave power (600–900 W) exerted no significant effect on hydrolysis. On enzymatic hydrolysis step,
improvement on solid percentage (16% w/v) and enzyme load (30 FPU gCAB-M−1) increased glucose concentration to 15 g L−1. The fermentation of the hydrolyzate by Saccharomyces cerevesiae resulted in ethanol concentration and productivity of 5.6 g L−1 and 1.41 g L−1 h−1, respectively. 相似文献
19.
A phosphite dehydrogenase gene (ptdhK) consisting of 1,011-bp nucleotides which encoding a peptide of 336 amino acid residues was cloned from Pseudomonas sp. K. gene ptdhK was expressed in Escherichia coli BL21 (DE3) and the corresponding recombinant enzyme was purified by metal affinity chromatography. The recombinant protein
is a homodimer with a monomeric molecular mass of 37.2 kDa. The specific activity of PTDH-K was 3.49 U mg−1 at 25 °C. The recombinant PTDH-K exhibited maximum activity at pH 3.0 and at 40 °C and displayed high stability within a
wide range of pHs (5.0 to 10.5). PTDH-K had a high affinity to its natural substrates, with K
m values for sodium phosphite and NAD of 0.475 ± 0.073 and 0.022 ± 0.007 mM, respectively. The activity of PTDH-K was enhanced
by Na+, NH4+, Mg2+, Fe2+, Fe3+, Co2+, and EDTA, and PTDH-K exhibited different tolerance to various organic solvents. 相似文献
20.
Bruno Benoliel Marcio José Poças-Fonseca Fernando Araripe Gonçalves Torres Lidia Maria Pepe de Moraes 《Applied biochemistry and biotechnology》2010,160(7):2036-2044
A β-glucosidase gene (bgl4) from Humicola grisea var thermoidea was successfully expressed in Saccharomyces cerevisiae. The recombinant protein (BGL4
Sc
) was initially detected associated with yeast cells and later in the culture medium. BGL4
Sc
showed optimal pH and temperature of 6.0 and 40 °C, respectively, and an apparent molecular mass of 57 kDa. The enzyme showed
activity against cellobiose and synthetic substrates, and was inhibited more than 80% by Fe2+, Cu2+, Zn2+, and Al3+. Using p-nitrophenyl-β-d-glucopyranoside (pNPG) as substrate, BGL4
Sc
presented a V
max of 6.72 μmol min−1 mg total protein−1 and a K
m of 0.16 mM under optimal conditions. Most important, BGL4
Sc
is resistant to inhibition by glucose and the calculated K
i value for this sugar is 70 mM. This feature prompts BLG4
Sc
as an ideal enzyme to be used in the saccharification process of lignocellulosic materials for ethanol production. 相似文献