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1.
Laila Manni Olfa Ghorbel-Bellaaj Kemel Jellouli Islem Younes Moncef Nasri 《Applied biochemistry and biotechnology》2010,162(2):345-357
Chitin is a polysaccharide found in abundance in the shell of crustaceans. In this study, the protease from Bacillus cereus SV1 was applied for chitin extraction from shrimp waste material of Metapenaeus monoceros. A high level of deproteinization 88.8% ± 0.4 was recorded with an E/S ratio of 20. The demineralization was completely achieved
within 6 h at room temperature in HCl 1.25 M, and the residual content of calcium in chitin was below 0.01%. 13C CP/MAS-NMR spectral analysis of chitin prepared by the enzymatic deproteinization of shrimp wastes was found to be similar
to that obtained by alkaline treatment and to the commercial α-chitin. The degree of N-acetylation, calculated from the spectrum,
was 89.5%. Chitin obtained by treatment with crude protease from B. cereus was converted to chitosan by N-deacetylation, and the antibacterial activity of chitosan solution against different bacteria
was investigated. Results showed that chitosan solution at 50 mg/mL markedly inhibited the growth of most Gram-negative and
Gram-positive bacteria tested. Furthermore, the antioxidant potential of the protein hydrolysates obtained during enzymatic
isolation of chitin was evaluated using various in vitro assays. All the samples exerted remarkable antioxidant activities. These results suggest that enzymatic deproteinization
of the shrimp shell wastes, using B. cereus SV1 protease, could be applicable to the chitin production process. 相似文献
2.
Ghorbel-Bellaaj O Jellouli K Younes I Manni L Ouled Salem M Nasri M 《Applied biochemistry and biotechnology》2011,164(4):410-425
A solvent-stable protease-producing bacterium was isolated and identified as Pseudomonas aeruginosa A2. The strain was found to produce high level of protease activity when grown in media containing only fresh shrimp waste
(FSW) or shrimp waste powder (SWP), indicating that it can obtain its carbon, nitrogen, and salts requirements directly from
shrimp waste. Maximum protease activities 17,000 and 12,000 U/mL were obtained with 80 g/L SWP and 135 g/L FSW, respectively.
The optimum temperature and pH for protease activity were 60 °C and 8.0, respectively. The crude protease, at different enzyme/substrate
(E/S) ratio, was tested for the deproteinization of shrimp waste to produce chitin. The crude enzyme of P. aeruginosa A2 was found to be effective in the deproteinization of shrimp waste. The protein removals after 3 h hydrolysis at 40 °C
with an E/S ratio of 0.5 and 5 U/mg protein were about 56% and 85%, respectively. 13C CP/MAS-NMR spectral analysis of the chitin prepared by treatment with the crude protease was carried out and was found to
be similar to that of the commercial α-chitin. These results suggest that enzymatic deproteinization of shrimp waste by A2
protease could be applicable to the chitin production process. 相似文献
3.
Ali Nel-H Hmidet N Ghorbel-Bellaaj O Fakhfakh-Zouari N Bougatef A Nasri M 《Applied biochemistry and biotechnology》2011,164(7):1096-1110
Alkaline proteases from the viscera of the striped seabream (Lithognathus mormyrus) were extracted and characterized. Interestingly, the crude enzyme was active over a wide range of pH from 6.0 to 11.0, with
an optimum pH at the range of 8.0–10.0. In addition, the crude protease was stable over a broad pH range (5.0–12.0). The optimum
temperature for enzyme activity was 50 °C. The crude alkaline proteases showed stability towards various surfactants and bleach
agents and compatibility with some commercial detergents. It was stable towards several organic solvents and retained more
than 50% of its original activity after 30 days of incubation at 30 °C in the presence of 25% (v/v) dimethyl sulfoxide, N,N-dimethylformamide, diethyl ether, and hexane. The crude enzyme extract was also tested for shrimp waste deproteinization
in the preparation of chitin. The protein removal with a ratio enzyme/substrate of 10 was about 79%. 相似文献
4.
A bacterial strain isolated from spoiled coconut and identified as Bacillus cereus was found capable of producing alkaline thermostable extracellular lipase. Optimum temperature, time, and pH for enzyme substrate
reaction were found to be 60 °C, 10 min, and 8.0 respectively. Common surfactants except Triton X 100 and cetyltrimethylammonium
bromide have no or very little inhibitory effects on enzyme activity. The enzyme was found to be stable in presence of oxidizing
agents and protease enzyme. The maximum lipase production was achieved at 30–33 °C, pH 8.0 on 24 h of fermentation using 50 ml
medium in a 250-ml Erlenmeyer flask. The superior carbon and nitrogen sources for lipase production were starch (2%) and ammonium
sulfate (nitrogen level 21.2 mg/100 ml), peptone (nitrogen level 297 mg/100 ml), and urea (nitrogen level 46.62 mg/100 ml)
in combination, respectively. The maximum enzyme activity obtained was 33 ± 0.567 IU/ml. 相似文献
5.
Chitinase was purified from the culture medium of Bacillus licheniformis SK-1 by colloidal chitin affinity adsorption followed by diethylamino ethanol-cellulose column chromatography. The purified
enzyme showed a single band on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The molecular size and pI of chitinase 72 (Chi72) were 72 kDa and 4.62 (Chi72) kDa, respectively. The purified chitinase revealed two activity optima
at pH 6 and 8 when colloidal chitin was used as substrate. The enzyme exhibited activity in broad temperature range, from
40 to 70°C, with optimum at 55°C. It was stable for 2 h at temperatures below 60°C and stable over a broad pH range of 4.0–9.0
for 24 h. The apparent K
m and V
max of Chi72 for colloidal chitin were 0.23 mg ml−1 and 7.03 U/mg, respectively. The chitinase activity was high on colloidal chitin, regenerated chitin, partially N-acetylated chitin, and chitosan. N-bromosuccinamide completely inhibited the enzyme activity. This enzyme should be a good candidate for applications in the
recycling of chitin waste. 相似文献
6.
Sequential acidic precipitation followed by a single chromatographic step (gel filtration) allowed the recovery of a collagenolytic
fraction containing several proteases from by-products of snow crab (Chionoecetes opilio). The partial purification was particularly efficient to recover tryptic (purification fold = 1,352.5; yield = 110%) but
also chymotryptic, elastolytic, and collagenolytic activities. A temperature of 40 °C and pH 8.0–8.5 were optimal for enzyme
activity, which was stable for 2 h under these conditions. Calcium was not required for stability and thus activity. The isoelectric
points of the protein components ranged from 3.7 to 4.6. Zymography revealed 29 and 48 kDa major components and others from
22 to 56 kDa. Enzymes were inhibited by PMSF and TLCK but were insensitive to TPCK. In view of these properties, the proteases
likely belong to the serine collagenase group. Inhibition by EDTA could be due to a mechanism other than Ca2+ chelation. Using a food system (ground fish), the fraction was more proteolytic than a commercial bacterial protease, suggesting
potential applications in enzymatic hydrolysis processes. 相似文献
7.
Mervat Morsy A. El-Gendy 《Applied biochemistry and biotechnology》2010,162(3):780-794
Among all endophytic keratinolytic fungal isolates recovered from marine soft coral Dendronephthya hemprichii, Penicillium spp. Morsy1 was selected as the hyperactive keratinolytic strain under solid substrate fermentation of different agriculture
and poultry wastes. The optimization of extraction process, physicochemical parameters affecting the keratinase production
in solid-state fermentation, and the purified keratinase parameters were studied. Maximum keratinase activity (1,600 U g−1, initial dry substrate) was recovered from moldy bran with 0.1% Tween 80. The optimized production conditions were rice straw
as carbon source, pH of medium 6, growth temperature 26 °C, initial moisture content of 80% (v/w), inoculum size of 105 spores ml−1, and an average particle size of the substrate 0.6 mm (3,560 U g−1, initial dry substrate after 5 days of fermentation). Two types of keratinase (Ahm1 and Ahm2) were purified from the culture
supernatant through ammonium sulfate precipitation, DEAE-Sepharose, and gel filtration chromatography. Enzyme molecular weights
were 19 kDa (Ahm1) and 40 kDa (Ahm2). The kinetic parameters of purified keratinases were optimized for the hydrolysis of
azokeratin by Ahm1 (pH 7.0–8.0, stable in pH range of 6.0 to 8.0 at 50 °C) and Ahm2 enzymes (pH 10.0–11.0, stable in pH range
of 6.0 to 11.0 at 60–65 °C). Whereas inhibitors of serine (phenylmethylsulfonyl fluoride) and cysteine (iodoacetamide) proteases
had minor effects on both Ahm1 and Ahm2 activity, both keratinases were strongly inhibited by chelating agents EDTA and EGTA.
These findings suggest that serine and cysteine residues are not involved in the catalytic mechanisms, and they are metalloproteases. 相似文献
8.
Yun Wang Jin-Zhu Song Qian Yang Zhi-Hua Liu Xiao-Mei Huang Yan Chen 《Applied biochemistry and biotechnology》2010,162(3):843-854
A gene encoding chitin deacetylase was cloned by polymerase chain reaction from Aspergillus nidulans. Sequencing result showed 40% homology to the corresponding gene from Colletotrichum lindemuthianum. The complete gene contains an open reading frame of 747 nucleotides encoding a sequence of 249 amino acid residues. The
chitin deacetylase gene was subcloned into a pET28a expression vector and expressed in Escherichia coli BL21 and then purified by metal affinity chromatography using a His-bind column. The purified chitin deacetylase demonstrated
an activity of 0.77 U ml−1 for the glycol chitin substrates, and its specific activity was 4.17 U mg−1 for it. The optimal temperature and pH of the purified enzyme were 50 °C and 8.0, respectively. When glycol chitin was used
as the substrate, K
m was 4.92 mg ml−1, and K
cat showed 6.25 s−1, thus the ratio of K
cat and K
m was 1.27 ml s−1 mg−1. The activity of chitin deacetylase was affected by a range of metal ions and ethylenediaminetetraacetic acid. 相似文献
9.
Qi Wu Chen Li Chenglei Li Hui Chen Liu Shuliang 《Applied biochemistry and biotechnology》2010,160(1):129-139
The collagenase, produced extracellular by Bacillus pumilus Col-J, was purified by ammonium sulfate precipitation followed by two gel filtrations, involving Sephadex G-100 column and
Sepharose Fast Flow column. Purified collagenase has a 31.53-fold increase in specific activity of 87.33 U/mg and 7.00% recovery.
The collagenase has a relative molecular weight of 58.64 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
The optimal temperature for the enzyme reaction was 45 °C. More than 50% of the original activity still remained after 5 min
of incubation at 70 °C or 10 min at 60 °C. The maximal enzyme activity of collagenase was obtained at pH 7.5, and it was stable
over a pH range of 6.5–8.0. The collagenase activity was strongly inhibited by Mn2+, Pb2+, ethylenediamine tetraacetic acid, ethylene glycol tetraacetic acid, and β-mercaptoethanol. However, Ca2+ and Mg2+ greatly increased its activity. The collagenase from B. pumilus Col-J showed highly specific activity towards the native collagen from calf skin. The K
m and V
max of the enzyme for collagen were 0.79 mg/mL and 129.5 U, respectively. 相似文献
10.
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
11.
Rym Agrebi Noomen Hmidet Mohamed Hajji Nawrez Ktari Anissa Haddar Nahed Fakhfakh-Zouari Moncef Nasri 《Applied biochemistry and biotechnology》2010,162(1):75-88
In this study, Mirabilis jalapa tuber powder (MJTP) was used as a new complex organic substrate for the growth and production of fibrinolytic enzymes by
a newly isolated Bacillus amyloliquefaciens An6. Maximum protease activity (1,057 U/ml) with casein as a substrate was obtained when the strain was grown in medium containing
(grams per liter) MJTP 30, yeast extract 6, CaCl2 1, K2HPO4 0.1, and K2HPO4 0.1. The strain was also found to grow and produce extracellular proteases in a medium containing only MJTP, indicating that
it can obtain its carbon, nitrogen, and salts requirements directly from MJTP. The B. amyloliquefaciens An6 fibrinase (BAF1) was partially purified, and fibrinolytic activity was assayed in a test tube with an artificial fibrin
clot. The molecular weight of the partially purified BAF1 fibrinolytic protease was estimated to be 30 kDa by sodium dodecyl
sulfate polyacrylamide gel electrophoresis and gel filtration. The optimum temperature and pH for the caseinolytic activity
were 60 °C and 9.0, respectively. The enzyme was highly stable from pH 6.0 to 11.0 and retained 62% of its initial activity
after 1 h incubation at 50 °C. However, the enzyme was inactivated at higher temperatures. The activity of the enzyme was
totally lost in the presence of phenylmethylsulfonyl fluoride, suggesting that BAF1 is a serine protease. 相似文献
12.
Phenylalanine dehydrogenase (l-PheDH) from Sporosarcina ureae was immobilized on DEAE-cellulose, modified initially with 2-amino-4,6-dichloro-s-triazine followed by hexamethylenediamine
and glutaraldehyde. The highest activity of immobilized PheDH was determined as 95.75 U/g support with 56% retained activity.
The optimum pH value of immobilized l-PheDH was shifted from pH 10.4 to 11.0. The immobilized l-PheDH showed activity variations close to the maximum value in a wider temperature range of 45–55 °C, whereas it was 40 °C
for the native enzyme. The pH and the thermal stability of the immobilized l-PheDH were also better than the native enzyme. At pH 10.4 and 25 °C, K
m values of the native and the immobilized l-PheDH were determined as K
m Phe = 0.118, 0.063 mM and K
m NAD+ = 0.234, 0.128 mM, respectively. Formed NADH at the exit of packed bed reactor column was detected by the flow-injection
analysis system. The conversion efficiency of the reactor was found to be 100% in the range of 5–600 μM Phe at 9 mM NAD+ with a total flow rate of 0.1 mL/min. The reactor was used for the analyses of 30 samples each for 3 h per day. The half-life
period of the reactor was 15 days. 相似文献
13.
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
14.
Daniela de Araújo Viana Carolina de Albuquerque Lima Rejane Pereira Neves Cristina Souza Mota Keila Aparecida Moreira José Luiz de Lima-Filho Maria Taciana Holanda Cavalcanti Attilio Converti Ana Lúcia Figueiredo Porto 《Applied biochemistry and biotechnology》2010,162(3):830-842
Cow raw milk from dairy cooperatives was examined for its microbial composition. Among the isolates identified, 17.6% were
yeasts. The most frequent genus was Candida, although members belonging to the genera Brettanomyces, Dekkera, and Geotricum were also identified. Although qualitative and quantitative tests for extracellular proteolytic activity were positive for
all the species isolated, Candida buinensis showed the highest response (23.5 U/mg); therefore, it was selected for subsequent investigation. The results of fermentations
carried out at variable temperature, pH, and soybean flour concentration, according to a 23 full factorial design, demonstrated that this yeast ensured the highest production of extracellular proteases (573 U/mL)
when cultivated at 35 °C, pH 6.5, and using soybean flour concentrations in the range 0.1–0.5% (w/v). The cell-free supernatants showed the highest activity at 25 °C and pH 7.0, and satisfactory stability in the ranges 25–30 °C
and pH 7–9. The first-order rate constants of protease inactivation in the cell-free supernatants were calculated at different
temperatures from semi-log plots of the residual activity versus time and then used in Arrhenius and Eyring plots to estimate
the main thermodynamic parameters of thermoinactivation (E* = 40.0 kJ/mol; ΔH* = 37.3 kJ/mol; ΔS* = −197.5 J/mol K; ΔG* = 101 kJ/mol). 相似文献
15.
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
16.
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
17.
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
18.
A heparinase-producing fungus was isolated, and the strain was taxonomically characterized as Aspergillus flavus by morphophysiological and 26S rRNA gene homology studies. The culture produced intracellular heparinase enzyme, which was
purified 40.5-fold by DEAE-Sephadex A-50, CM-Sephadex C-50, and Sephadex G-100 column chromatography. Specific activity of
the purified enzyme was found to be 44.6 IU/μg protein and the molecular weight of native as well as reduced heparinase was
24 kDa, showing a monomeric unit structure. Peptide mass spectrum showed poor homogeneity with the database in the peptide
bank. The enzyme activity was maximum at 30 °C in the presence of 300 mM NaCl at pH 7.0. In the presence of Co2+, Mn2+ ions, and reducing agents (β-mercaptoethanol, dithiothreitol), enzyme activity was enhanced and inhibited by iodoacetic acid.
These observations suggested that free sulfohydryl groups of cysteine residues were necessary for catalytic activity of the
enzyme. The enzyme was also inhibited by histidine modifier, DEPC, which suggests that along with cysteine, histidine may
be present at its active site. The enzyme showed a high affinity for heparin as a substrate with K
m and V
max as 2.2 × 10−5 M and 30.8 mM min−1, respectively. The affinity of the enzyme for different glycosaminoglycans studied varied, with high substrate specificity
toward heparin and heparin-derived polysaccharides. Depolymerization of heparin and fractionation of the oligosaccharides
yielded heparin disaccharides as main product. 相似文献
19.
Purification,Properties, and Application of a Novel Acid Urease from <Emphasis Type="Italic">Enterobacter</Emphasis> sp. 总被引:1,自引:0,他引:1
It has been demonstrated that acid urease is capable of decomposing urea in fermented beverage and foods. As urea is a precursor
of ethylcarbamate, a potential carcinogenic compound, measures must be taken to control the level of urea. We herein describe
the purification and characterization of a novel acid urease from Enterobacter sp. R-SYB082 and its application to the removal of urea in Chinese rice wine. The enzyme was purified to electrophoretic
homogeneity using ethanol precipitation, Superdex 200 and Mono Q with a fold purification of 21.1 and a recovery of 49%. The
molecular weight of the enzyme was 430,000 Da by gel filtration and 72,000 Da by sodium dodecyl sulfate polyacrylamide gel
electrophoresis, suggesting that it was a hexamer. The activity of this purified enzyme was optimal at pH 4.5 and 35 °C. The
temperature stability was under 55 °C, and the pH stability was 4.0~5.0. The enzyme exhibited an apparent K
m of 19.5 μmol/l and a V
max of 109 μmol urea/mg·min at 35 °C and pH 4.5. When incubating two different kinds of Chinese rice wine with the enzyme (0.08 U/ml)
at 35 °C for 7 days, over 85% of urea was decomposed, and at 20 °C, above 78% was removed. The result showed that the enzyme
is applicable to elimination of urea in Chinese rice wine. 相似文献
20.
A 56.56-kDa extracellular chitinase from Paenibacillus sp. D1 was purified to 52.3-fold by ion exchange chromatography using SP Sepharose. Maximum enzyme activity was recorded
at pH 5.0 and 50 °C. MALDI-LC-MS/MS analysis identified the purified enzyme as chitinase with 60% similarity to chitinase
Chi55 of Paenibacillus ehimensis. The activation energy (E
a) for chitin hydrolysis and temperature quotient (Q
10) at optimum temperature was found to be 19.14 kJ/mol and 1.25, respectively. Determination of kinetic constants k
m, V
max, k
cat, and k
cat/k
m and thermodynamic parameters ΔH*, ΔS*, ΔG*, ΔG*E–S, and ΔG*E–T revealed high affinity of the enzyme for chitin. The enzyme exhibited higher stability in presence of commonly used protectant
fungicides Captan, Carbendazim, and Mancozeb compared to control as reflected from the t
1/2 values suggesting its applicability in integrated pest management for control of soil-borne fungal phytopathogens. The order
of stability of chitinase in presence of fungicides at 80 °C as revealed from t
1/2 values and thermodynamic parameters E
a(d) (activation energy for irreversible deactivation), ΔH*, ΔG*, and ΔS* was: Captan > Carbendazim > Mancozeb > control. The present study is the first report on thermodynamic and kinetic characterization
of chitinase from Paenibacillus sp. D1. 相似文献