共查询到20条相似文献,搜索用时 15 毫秒
1.
Chellappan S Jasmin C Basheer SM Kishore A Elyas KK Bhat SG Chandrasekaran M 《Journal of industrial microbiology & biotechnology》2011,38(6):743-752
An alkaline protease from marine Engyodontium album was characterized for its physicochemical properties towards evaluation of its suitability for potential industrial applications.
Molecular mass of the enzyme by matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS) analysis was calculated
as 28.6 kDa. Isoelectric focusing yielded pI of 3–4. Enzyme inhibition by phenylmethylsulfonyl fluoride (PMSF) and aprotinin
confirmed the serine protease nature of the enzyme. K
m, V
max, and K
cat of the enzyme were 4.727 × 10−2 mg/ml, 394.68 U, and 4.2175 × 10−2 s−1, respectively. Enzyme was noted to be active over a broad range of pH (6–12) and temperature (15–65°C), with maximum activity
at pH 11 and 60°C. CaCl2 (1 mM), starch (1%), and sucrose (1%) imparted thermal stability at 65°C. Hg2+, Cu2+, Fe3+, Zn2+, Cd+, and Al3+ inhibited enzyme activity, while 1 mM Co2+ enhanced enzyme activity. Reducing agents enhanced enzyme activity at lower concentrations. The enzyme showed considerable
storage stability, and retained its activity in the presence of hydrocarbons, natural oils, surfactants, and most of the organic
solvents tested. Results indicate that the marine protease holds potential for use in the detergent industry and for varied
applications. 相似文献
2.
A highly selective sucrose isomerase (SIase) was purified to homogeneity from the cell-free extract of Erwinia rhapontici NX-5 with a recovery of 27.7% and a fold purification of 213.6. The purified SIase showed a high specific activity of 427.1 U mg−1 with molecular weight of 65.6 kDa. The K
m for sucrose was 222 mM while V
max was 546 U mg−1. The optimum pH and temperature for SIase activity were 6.0 and 30 °C, respectively. The purified SIase was stable in the
temperature range of 10–40 °C and retained 65% of the enzyme activity after 2 weeks’ storage at 30 °C. The SIase activity
was enhanced by Mg2+ and Mn2+, inhibited by Ca2+, Cu2+, Zn2+, and Co2+, completely inhibited by Hg2+ and Ag2+. The purified SIase was strongly inhibited by SDS, while partially inhibited by dimethylformamide, tetrahydrofuran, and PMSF.
Additionally, glucose and fructose acted as competitive inhibitors for purified SIase. 相似文献
3.
The invertase of Lactobacillus reuteri CRL 1100 is a glycoprotein composed by a single subunit with a molecular weight of 58 kDa. The enzyme was stable below 45°C
over a wide pH range (4.5–7.0) with maximum activity at pH 6.0 and 37°C. The invertase activity was significantly inhibited
by bivalent metal ions (Ca++, Cu++, Cd++, and Hg++), β-mercaptoethanol, and dithiothreitol and partially improved by ethylenediaminetetraacetic acid. The enzyme was purified
32 times over the crude extract by gel filtration and ion-exchange chromatography with a recovery of 17%. The K
m
and Vmax values for sucrose were 6.66 mM and 0.028 μmol/min, respectively. An invertase is purified and characterized for the first
time in Lactobacillus, and it proved to be a β-fructofuranosidase.
Received: 13 August 1999 / Accepted: 15 September 1999 相似文献
4.
Laura Dipasquale Agata Gambacorta Rosa Anna Siciliano Maria Fiorella Mazzeo Licia Lama 《Extremophiles : life under extreme conditions》2009,13(2):345-354
This is the first report describing the purification and enzymatic properties of a native invertase (β-D-fructosidase) in
Thermotogales. The invertase of the hydrogen-producing thermophilic bacterium Thermotoga
neapolitana DSM 4359 (hereby named Tni) was a monomer of about 47 kDa having an amino acid sequence quite different from other invertases studied up to now. Its
properties and substrates specificity let us classify this protein as a solute-binding protein with invertase activity. Tni was specific for the fructose moiety and the enzyme released fructose from sucrose and raffinose and the fructose polymer
inulin was hydrolyzed in an endo-type fashion. Tni had an optimum temperature of 85°C at pH 6.0. At temperatures of 80–85°C, the enzyme retained at least 50% of its initial
activity during a 6 h preincubation period. Tni had a K
m
and k
cat
/K
m
values (at 85°C and pH 6.0) of about 14 mM and 5.2 × 108 M−1 s−1, respectively.
Dedicated to the memory of Prof. R. A. Nicolaus, founder of the Institute (1968). 相似文献
5.
Effect of salicylic and abscisic acid administered through detached tillers on antioxidant system in developing wheat grains under heat stress 总被引:1,自引:0,他引:1
The mechanism imparting thermotolerance by salicylic acid (SA) and abscisic acid (ABA) is still unresolved using either spraying
technique or in vitro conditions. Alternative way of studying these effects under near in vivo conditions is through the use
of liquid culturing technique. Effects of SA and ABA (100 μM) on antioxidative enzymes, antioxidants and lipid peroxidation
were studied in detached tillers of three wheat (Triticum aestivum L.) cultivars PBW 343, C 306 (heat tolerant) and WH 542 (heat susceptible) cultured in a liquid medium. Ears were subjected
to heat shock treatment (45°C for 2 h) and then maintained at 25°C for 5 days. Heat shock treatment resulted in increased
peroxidase (POD) activity, while superoxide dismutase (SOD) and catalase (CAT) activities were reduced compared to control.
The decrease in CAT activity was more significant in susceptible cultivar WH 542. Concomitantly, content of α-tocopherol and
lipid peroxides increased in heat-treated wheat ears, whereas contents of total ascorbate level were reduced. Following treatment
with SA and ABA, activities of all three antioxidative enzymes increased in correspondence with an increase in ascorbate and
α-tocopherol content. Apparently, lipid peroxide content was reduced by SA in heat tolerant cultivars (PBW 343 and C 306)
whereas in susceptible cultivar it was decreased by ABA. The up-regulation of the antioxidant system by SA and ABA possibly
contributes to better tolerance against heat shock-induced oxidative damage in wheat grains. 相似文献
6.
The present study investigates the efficiency of Aspergillus niger to produce invertase, an industrially important enzyme by using powdered stem of Cympopogan caecius (Lemon grass) as sole substrate and sole carbon source for the microorganism. The molecular weight of invertase was estimated
to be 66–70 kDa by sodium do decyl sulphate poly acrylamide gel electrophoresis (SDS PAGE). The production of the enzyme was
studied at different pH scales ranging from pH 4.0 to 7.0 at a constant temperature of 30°C and 2% substrate concentration.
The maximum production of invertase (specific activity −0.0516 μk/mg protein) was obtained at pH 5.5 at 30°C temperature,
and incubation for 48 h. The activity was found to be stable at pH 5.5 for 30 min. The enzyme was found to be stable in the
temperature range of 20–55°C. The effect of divalent metal ions Cu2+, Fe2+, Co2+ on the activity of the enzyme invertase showed that these ions affected the activity by a certain factor. The study can be
further industrially exploited in a country-like India where lemon grass is found in plenty and can be used as substrate for
enzyme production. Moreover, the preparation of the substrate is also a simple process. 相似文献
7.
Pratibha Dheeran Sachin Kumar Yogesh K. Jaiswal Dilip K. Adhikari 《Applied microbiology and biotechnology》2010,86(6):1857-1866
A newly isolated Geobacillus sp. IIPTN (MTCC 5319) from the hot spring of Uttarakhand's Himalayan region produced a hyperthermostable α-amylase. The microorganism
was characterized by biochemical tests and 16S rRNA gene sequencing. The optimal temperature and pH were 60°C and 6.5, respectively,
for growth and enzyme production. Although it was able to grow in temperature ranges from 50 to 80°C and pH 5.5–8.5. Maximum
enzyme production was in exponential phase with activity 135 U ml−1 at 60°C. Assayed with cassava as substrate, the enzyme displayed optimal activity 192 U ml−1 at pH 5.0 and 80°C. The enzyme was purified to homogeneity with purification fold 82 and specific activity 1,200 U mg−1 protein. The molecular mass of the purified enzyme was 97 KDa. The values of K
m
and V
max were 36 mg ml−1 and 222 μmol mg−1 protein min−1, respectively. The amylase was stable over a broad range of temperature from 40°C to 120°C and pH ranges from 5 to 10. The
enzyme was stimulated with Mn2+, whereas it was inhibited by Hg2+, Cu2+, Zn2+, Mg2+, and EDTA, suggesting that it is a metalloenzyme. Besides hyperthermostability, the novelty of this enzyme is resistance
against protease. 相似文献
8.
Gulelat D. Haki Alfredo J. Anceno Sudip K. Rakshit 《World journal of microbiology & biotechnology》2008,24(11):2517-2524
Bacillus sp. GRE1 isolated from an Ethiopian hyperthermal spring produced raw-starch digesting, Ca2+-independent thermostable α-amylase. Enzyme production in shake flask experiments using optimum nutrient supplements and environmental
conditions was 2,360 U l−1. Gel filtration chromatography yielded a purification factor of 33.6-fold and a recovery of 46.5%. The apparent molecular
weight of the enzyme was 55 kDa as determined by SDS-PAGE. Presence or absence of Ca2+ produced similar temperature optima of 65–70°C. The optimum pH was in the range of 5.5–6.0. The enzyme maintained 50% of
its original activity after 45 min of incubation at 80°C and was stable at a pH range of 5.0–9.0. The V
max and K
m values for soluble starch were 42 mg reducing sugar min−1 and 4.98 mg starch ml−1, respectively. Strong inhibitors of enzyme activity included Cu2+, Zn2+ and Fe2+. The enzyme coding gene and the deduced protein translation revealed a characteristic but markedly atypical homology to Bacillus species α-amylase sequences. The enzyme hydrolyzed wheat, corn and tapioca starch granules efficiently below their gelatinization
temperatures. Rather than the higher oligosaccharides normally produced by Bacillus α-amylases operating at high temperatures, maltose was the major hydrolysis product with the present enzyme. 相似文献
9.
Xylaria regalis, a wood-grown ascomycete isolated in Taiwan, produces β-glucosidase (EC 3.2.1.21) extracellularly. The β-glucosidase was
purified to homogeneity by ammonium sulfate precipitation, ion-exchange, and gel filtration chromatography. The molecular
mass of the purified enzyme was estimated to be 85 kDa by sodium dodecyl sulfate–polyacrylamide gel electrophoresis. With
p-nitrophenyl β-D-glucopyranoside (PNPG) as the substrate at pH 5.0 and 50°C, the K
m
was 1.72 mM and V
max
was 326 μmol/min/mg. Optimal activity with PNPG as the substrate was at pH 5.0 and 50°C. The enzyme was stable at pH 5.0
at temperatures up to 50°C. The purified β-glucosidase was active against PNPG, cellobiose, sophorose, and gentiobiose, but
did not hydrolyze lactose, sucrose, Avicel, and o-nitrophenyl β-D-galactopyranoside. The activity of β-glucosidase was stimulated by Ca2+, Mg2+, Mn2+, Cd2+ and β-mercaptoethanol, and inhibited by Ag+, Hg2+, SDS, and p-chloromercuribenzoate (PCMB).
Received: 30 March 1996 / Accepted: 3 May 1996 相似文献
10.
Guo Li-Qiong Lin Shuo-Xin Zheng Xiao-Bing Huang Zi-Rou Lin Jun-Fang 《World journal of microbiology & biotechnology》2011,27(3):731-735
A thermostable laccase was isolated from a tropical white-rot fungus Polyporus sp. which produced as high as 69,738 units of laccase l−1 in an optimized medium containing 20 g of malt extract l−1, 2 g of yeast extract l−1, 1.5 mM CuSO4. The laccase was purified to electrophoretic purity with a final purification of 44.70-fold and a recovery yield of 21.04%.
The purified laccase was shown to be a monomeric enzyme with a molecular mass of 60 kDa. The optimum temperature and pH value
of the laccase were 75°C and pH 4.0, respectively, for 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonate) (ABTS). The Michaelis–Menten
constant (K
m
) of the laccase was 18 μM for ABTS substrate. The laccase was stable at pH values between 5.5 and 7.5. About 80% of the initial
enzyme activity was retained after incubation of the laccase at 70°C for 2 h, indicating that the laccase was intrinsically
highly thermostable and with valuable potential applications. The laccase activity was promoted by 4.0 mM of Mg2+, Mn2+, Zn2+ and Ca2+, while inhibited by 4.0 mM of Co2+, Al3+, Cu2+, and Fe2+, showing different profiles of metal ion effects. 相似文献
11.
A cold-adapted α-amylase (ParAmy) gene from Pseudoalteromonas
arctica GS230 was cloned, sequenced, and expressed as an N-terminus His-tag fusion protein in E. coli. A recombinant protein was produced and purified with DEAE-sepherose ion exchange chromatography and Ni affinity chromatography.
The molecular weight of ParAmy was estimated to be 55 KDa with sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE). With an optimum
temperature for activity 30 °C, ParAmy showed 34.5% of maximum activity at 0 °C and its activity decreased sharply at above 40 °C. ParAmy was stable in the range of pH 7–8.5 at 30 °C for 1 h. ParAmy was activated by Mn2+, K+ and Na+, and inhibited by Hg2+, Cu2+, and Fe3+. N-Bromosuccinimid showed a significant repressive effect on enzyme activity. The K
m and V
max values of the α-amylase for soluble starch were 7.28 mg/mL and 13.07 mg/mL min, respectively. This research suggests that
Paramy has a good potential to be a cold-stable and alkalitolerant amylase in detergent industry. 相似文献
12.
M Saleem F Aslam MS Akhtar M Tariq MI Rajoka 《World journal of microbiology & biotechnology》2012,28(2):513-522
Delignification efficacy of xylanases to facilitate the consequent chemical bleaching of Kraft pulps has been studied widely.
In this work, an alkaline and thermally stable cellulase-less xylanase, derived from a xylanolytic Bacillus subtilis, has been purified by a combination of gel filtration and Q-Sepharose chromatography to its homogeneity. Molecular weight
of the purified xylanase was 61 kDa by SDS–PAGE. The purified enzyme revealed an optimum assay temperature and pH of 60°C
and 8.0, respectively. Xylanase was active in the pH range of 6.0–9.0 and stable up to 70°C. Divalent ions like Ca2+, Mg2+ and Zn2+ enhanced xylanase activity, whereas Hg2+, Fe2+, and Cu2+ were inhibitory to xylanase at 2 mM concentration. It showed K
m
and V
max
values of 9.5 mg/ml and 53.6 μmol/ml/min, respectively, using birchwood xylan as a substrate. Xylanase exhibited higher values
of turn over number (K
cat) and catalytic efficiency (K
cat/K
m) with birchwood xylan than oat spelt xylan. Bleach-boosting enzyme activity at 30 U/g dry pulp displayed the optimum bio-delignification
of Kraft pulp resulting in 26.5% reduction in kappa number and 18.5% ISO induction in brightness at 55°C after 3 h treatment.
The same treatment improved the pulp properties including tensile strength and burst index, demonstrating its potential application
in pre-bleaching of Kraft pulp. 相似文献
13.
Verónica Beatriz Rajal Alicia Graciela Cid Guillermo Ellenrieder Carlos Mario Cuevas 《World journal of microbiology & biotechnology》2009,25(6):1025-1033
Penicillium ulaiense is a post-harvest pathogenic fungus that attacks citrus fruits. The objective of this work was to study this microorganism
as an α-l-rhamnosidase producer and to characterize it from P. ulaiense. The enzyme under study is used for different applications in food and beverage industries. α-l-Rhamnosidase was produced in a stirred-batch reactor using rhamnose as the main carbon source. The kinetic parameters for
the growth of the fungi and for the enzyme production were calculated from the experimental values. A method for partial purification,
including (NH4)2SO4 precipitation, incubation at pH 12 and DEAE-sepharose chromatography yielded an enzyme with very low β-glucosidase activity.
The pH and temperature optima were 5.0 and 60°C, respectively. The Michaelis–Menten constants for the hydrolysis of p-nitrophenyl-α-l-rhamnoside were V
max = 26 ± 4 IU ml−1 and K
m
= 11 ± 2 mM. The enzyme showed good thermostability up to 60°C and good operational stability in white wine. Co2+ affected positively the activity; EDTA, Mn2+, Mg2+, dithiotreitol and Cu2+ reduced the activity by different amounts, and Hg2+ completely inhibited the enzyme. The enzyme showed more activity on p-nitrophenyl-α-l-rhamnoside than on naringin. According to these results, this enzyme has potential for use in the food and pharmacy industries
since P. ulaiense does not produce mycotoxins. 相似文献
14.
Inhibition of ribonuclease and protease activities in germinating rice seeds exposed to nickel 总被引:1,自引:0,他引:1
When the seeds of two rice cvs. Malviya-36 and Pant-12 were germinated up to 120 h in the presence of 200 and 400 μM NiSO4, a significant reduction in the germination of seeds occurred. Seeds germinating in the presence of 400 μM NiSO4 showed about 12–20% decline in germination percent, about 20–53% decline in lengths and about 8–34% decline in fresh weights
of roots and shoots at 120 h of germination. Ni2+ exposure of germinating seeds resulted in apparent increased levels of RNA, soluble proteins, and free amino acids in endosperms
as well as embryo axes. A 400 μM Ni2+ treatment led to about 58–101% increase in the level of soluble proteins and about 39–107% increase in the level of free
amino acids in embryo axes at 96 h of germination. Activities of ribonuclease and protease declined significantly with increasing
levels of Ni2+ treatment. Isoenzyme profile of RNase as revealed by activity staining indicated decline in the intensities of 3–4 preexisting
enzyme isoforms in embryo axes of both the rice cultivars and disappearance of one of the two isoforms in endosperms of cv. Pant-12 due to 400 μM Ni2+ treatment. Results suggest that the presence of high level of Ni2+ in the medium of germinating rice seeds serves as a stress factor resulting in decreased hydrolysis as well as delayed mobilization
of endospermic RNA and protein reserves and causing imbalance in the level of biomolecules like RNA, proteins, and amino acids
in growing embryo axes. These events would ultimately contribute to decreased germination of rice seeds in high Ni2+ containing environment. 相似文献
15.
Yuzbasheva EY Yuzbashev TV Laptev IA Konstantinova TK Sineoky SP 《Applied microbiology and biotechnology》2011,89(3):645-654
The gene encoding homodimeric β-galactosidase (lacA) from Bacillus licheniformis DSM 13 was cloned and overexpressed in Escherichia coli, and the resulting recombinant enzyme was characterized in detail. The optimum temperature and pH of the enzyme, for both
o-nitrophenyl-β-d-galactoside (oNPG) and lactose hydrolysis, were 50°C and 6.5, respectively. The recombinant enzyme is stable in the range of pH 5 to 9 at
37°C and over a wide range of temperatures (4–42°C) at pH 6.5 for up to 1 month. The K
m values of LacA for lactose and oNPG are 169 and 13.7 mM, respectively, and it is strongly inhibited by the hydrolysis products, i.e., glucose and galactose.
The monovalent ions Na+ and K+ in the concentration range of 1–100 mM as well as the divalent metal cations Mg2+, Mn2+, and Ca2+ at a concentration of 1 mM slightly activate enzyme activity. This enzyme can be beneficial for application in lactose hydrolysis
especially at elevated temperatures due to its pronounced temperature stability; however, the transgalactosylation potential
of this enzyme for the production of galacto-oligosaccharides (GOS) from lactose was low, with only 12% GOS (w/w) of total sugars obtained when the initial lactose concentration was 200 g/L. 相似文献
16.
We have identified two types of peroxidases (POX), one ionically and one covalently bound to the particulate fraction, in
stripe rust-infected and -uninfected wheat (Triticum aestivum L.) leaves. The cell walls contained a high level of POX, of which 73–76% was extractable by 1% NaCl and 24–26% by 5 mM EDTA in infected and non-infected leaves of HD 2329. The NaCl-released POX constituted the predominant fraction. Both NaCl-
and EDTA-extracted POX exhibited maximum activity at pH 5.0 and had a K
m (enzyme–substrate affinity measure) value of 1.61–1.70 and 1.64–1.67 mM, respectively, with o-dianisidine as the substrate. The V
max (maximum catalytic rate) in the two extractions ranged between 7.06–7.45 and 6.65–7.82 μmol min−1 g−1 fresh weight. A temperature optimum of 50°C was observed for both the NaCl- and EDTA-released fractions. The two POX fractions
showed a differential response to metal ions, suggesting their distinctive nature. Sodium azide inhibited POX activity markedly,
which suggested the presence of heme as a prosthetic group. Inhibition of wall-bound POX by iodine and the regeneration of
activity by mercaptoethanol suggested the involvement of cysteine in the active site of the enzyme. These two forms showed
greater differences in terms of thermodynamic properties, such as the energy of activation (E
a) and enthalpy change (ΔH), while entropy (ΔS) and free energy changes were similar. The results further show that pathogen infection of the leaves of this susceptible
wheat cultivar induces an increase in the activity and kinetics of POX, which may be critical in the response of the plant
cell to infection. 相似文献
17.
A β-mannanase gene, designated as man5S27, was cloned from Streptomyces sp. S27 using the colony polymerase chain reaction (PCR) method and expressed in Escherichia coli BL21 (DE3). The open reading frame consisted of 1,161 bp and encoded a 386-amino-acid polypeptide (Man5S27) with calculated
molecular mass of 37.2 kDa. The encoded protein comprised a putative 38-residue signal peptide, a family 5 glycoside hydrolase
domain, and a family 10 carbohydrate-binding module. Purified recombinant Man5S27 had high specific activity of 2,107 U mg−1 and showed optimal activity at pH 7.0 and 65°C. The enzyme remained stable at pH 5.0–9.0 and had good thermostability at
50°C. The K
m values for locust bean gum and konjac flour were 0.16 and 0.41 mg ml−1, with V
max values of 3,739 and 1,653 μmol min−1 mg−1, respectively. Divalent metal ions such as Mn2+, Zn2+, Ca2+, Pb2+, and Fe2+ enhanced the enzyme activity, but Ag+ and Hg2+ strongly inhibited the activity. Man5S27 also showed resistance to various neutral proteases (retaining >95% activity after
proteolytic treatment for 2 h). 相似文献
18.
MA Giraldo TM da Silva F Salvato HF Terenzi JA Jorge LH Guimarães 《World journal of microbiology & biotechnology》2012,28(2):463-472
The filamentous fungus Paecylomices variotii was able to produce high levels of cell extract and extracellular invertases when grown under submerged fermentation (SbmF)
and solid-state fermentation, using agroindustrial products or residues as substrates, mainly soy bran and wheat bran, at
40°C for 72 h and 96 h, respectively. Addition of glucose or fructose (≥1%; w/v) in SbmF inhibited enzyme production, while
the addition of 1% (w/v) peptone as organic nitrogen source enhanced the production by 3.7-fold. However, 1% (w/v) (NH4)2HPO4 inhibited enzyme production around 80%. The extracellular form was purified until electrophoretic homogeneity (10.5-fold
with 33% recovery) by DEAE-Fractogel and Sephacryl S-200 chromatography. The enzyme is a monomer with molecular mass of 102 kDa
estimated by SDS–PAGE with carbohydrate content of 53.6%. Optima of temperature and pH for both, extracellular and cell extract
invertases, were 60°C and 4.0–4.5, respectively. Both invertases were stable for 1 h at 60°C with half-lives of 10 min at
70°C. Mg2+, Ba2+ and Mn2+ activated both extracellular and cell extract invertases from P. variotii. The kinetic parameters Km and Vmax for the purified extracellular enzyme corresponded to 2.5 mM and 481 U/mg prot−1, respectively. 相似文献
19.
Asiya Nazir Rohit Soni H. S. Saini R. K. Manhas B. S. Chadha 《World journal of microbiology & biotechnology》2009,25(7):1189-1197
An endoglucanase (1, 4-β-d glucan glucanohydrolase, EC 3.2.1.4) which was catalytically more active and exhibited higher affinity towards barley β-glucan,
xyloglucan and lichenin as compared to carboxymethylcellulose (CMC) was purified from Aspergillus terreus strain AN1 following ion-exchange and hydrophobic interaction chromatography and gel filtration. The purified enzyme (40-fold) that
apparently lacked a cellulose-binding domain showed a specific activity of 60 μmol mg−1 protein−1 against CMC. The purified enzyme had a molecular weight of 78 and 80 KDa as indicated by sodium dodecyl sulphate–polyacrylamide
gel electrophoresis and gel filtration, respectively, and a pI of 3.5. The enzyme was optimally active at temperature 60°C
and pH 4.0, and was stable over a broad range of pH (3.0–5.0) at 50°C. The endoglucanase activity was positively modulated
in the presence of Cu2+, Mg2+, Ca2+, Na+, DTT and mercaptoethanol. Endoglucanase exhibited maximal turn over number (K
cat) and catalytic efficiency (K
cat/km) of 19.11 × 105 min−1 and 29.7 × 105 mM−1 min−1 against barley β-glucan as substrate, respectively. Hydrolysis of CMC and barley β-glucan liberated cellobiose, cellotriose,
cellotetraose and detectable amount of glucose. The hydrolysis of xyloglucan, however, apparently yielded positional isomers
of cellobiose, cellotriose and cellotetraose as well as larger oligosaccharides. 相似文献
20.
Rojas NL Voget CE Hours RA Cavalitto SF 《Journal of industrial microbiology & biotechnology》2011,38(9):1515-1522
Rhamnosidases are enzymes that catalyze the hydrolysis of terminal nonreducing L-rhamnose for the bioconversion of natural or synthetic rhamnosides. They are of great significance in the current biotechnological
area, with applications in food and pharmaceutical industrial processes. In this study we isolated and characterized a novel
alkaline rhamnosidase from Acrostalagmus luteo albus, an alkali-tolerant soil fungus from Argentina. We also present an efficient, simple, and inexpensive method for purifying
the A. luteo albus rhamnosidase and describe the characteristics of the purified enzyme. In the presence of rhamnose as the sole carbon source,
this fungus produces a rhamnosidase with a molecular weight of 109 kDa and a pI value of 4.6, as determined by SDS–PAGE and
analytical isoelectric focusing, respectively. This enzyme was purified to homogeneity by chromatographic and electrophoretic
techniques. Using p-nitrofenil-α-L-rhamnopiranoside as substrate, the enzyme activity showed pH and temperature optima of 8.0 and 55°C, respectively. The enzyme
exhibited Michaelis–Menten kinetics, with K
M and V
max values of 3.38 mmol l−1 and 68.5 mmol l−1 min−1, respectively. Neither divalent cations such as Ca2+, Mg2+, Mn2+, and Co2+ nor reducing agents such as β-mercaptoethanol and dithiothreitol showed any effect on enzyme activity, whereas this activity
was completely inhibited by Zn2+ at a concentration of 0.2 mM. This enzyme showed the capacity to hydrolyze some natural rhamnoglucosides such as hesperidin,
naringin and quercitrin under alkaline conditions. Based on these results, and mainly due to the high activity of the A. luteo albus rhamnosidase under alkaline conditions, this enzyme should be considered a potential new biocatalyst for industrial applications. 相似文献