A cellulase producing bacterium (E3 strain) was isolated from fecal matter of elephant and identified as Bacillus sp. using 16S rDNA sequenced based molecular phylogenetic approach. While studying the effect of substr...A cellulase producing bacterium (E3 strain) was isolated from fecal matter of elephant and identified as Bacillus sp. using 16S rDNA sequenced based molecular phylogenetic approach. While studying the effect of substrates like Carboxymethyl cellulose (CMC), avicel, starch, maltose, sucrose, glucose, fructose, galactose and lactose on cellulase production, it was found that CMC was best carbon source induced cellulase production followed by lactose in this bacterial strain. A positive synergistic effect of lactose with CMC was also observed with enhancement of 5 - 6 times in cellulase production. The optimum cellulase production was recorded with 1% CMC and 1% lactose when added individually in the Omeliansky’s medium. The results showed that addition lactose with CMC greatly enhances the production and activity of various cellulase enzymes. The optimal fermentation conditions for the biosynthesis of cellulase by this strain were found to be temperature: 37℃, pH 7.0. The nitrogen source NH4Cl at 0.15% was optimum for cellulase production by this bacterium.展开更多
We have reported that A. pullulans 98 produces a high yield of cellulase. In this study, a carboxymethyl cellulase(CMCase) in the supernatant of the culture of A. pullulans 98 was purified to homogeneity, and the maxi...We have reported that A. pullulans 98 produces a high yield of cellulase. In this study, a carboxymethyl cellulase(CMCase) in the supernatant of the culture of A. pullulans 98 was purified to homogeneity, and the maximum production of CMCase was 4.51 U(mg protein)-1. The SDS-PAGE analysis showed that the molecular mass of the purified CMCase was 67.0 k Da. The optimal temperature of the purified enzyme with considerable thermosensitivity was 40℃, much lower than that of the CMCases from other fungi. The optimal p H of the enzyme was 5.6, and the activity profile was stable in a range of acidity(p H 5.0–6.0). The enzyme was activated by Na+, Mg2+, Ca2+, K+, Fe2+ and Cu2+, however, it was inhibited by Fe3+, Ba2+, Zn2+, Mn2+ and Ag+. Km and Vmax values of the purified enzyme were 4.7 mg m L-1 and 0.57 μmol L-1 min-1(mg protein)-1, respectively. Only oligosaccharides with different sizes were released from carboxymethylcellulose(CMC) after hydrolysis with the purified CMCase. The putative gene encoding CMCase was cloned from A. pullulans 98, which contained an open reading frame of 954 bp(EU978473). The protein deduced contained the conserved domain of cellulase superfamily(glucosyl hydrolase family 5). The N-terminal amino acid sequence of the purified CMCase was M-A-P-H-A-E-P-Q-S-Q-T-T-E-Q-T-S-S-G-Q-F, which was consistent with that deduced from the cloned gene. This suggested that the purified CMCase was indeed encoded by the cloned CMCase gene in this yeast.展开更多
Five thermophilic strains that can degrade cellulose were isolated from the compost of a waste management in Guangzhou, China. Since one of them degraded cellulose effectively, it was chosen as the study strain. Based...Five thermophilic strains that can degrade cellulose were isolated from the compost of a waste management in Guangzhou, China. Since one of them degraded cellulose effectively, it was chosen as the study strain. Based on its morphology, spores′ susceptibility to heat, cell wall composition and other characteristics, the organism was classified as Thermomonospora fusca. Conditions for production of carboxy methyl cellulase (CMCase) were examined. The optimal temperature and pH value for enzyme production were 50 ℃ and 10.5, respectively. Cellulosic materials and easily metabolisable carbohydrates served as carbon sources for the growth of the strain. Only cotton, avicel,carboxy methyl cellulose (CMC) acted as potent inducers for the production of cellulases by this strain. Despite excellent growth on easily metabolisable carbohydrates, only constitutive levels of cellulases were produced. The optimal carbon and nitrogen sources for CMCase production were cotton and soybean respectively. The high thermostability, wide pH stability, and cheap nitrogen source show well potential use for composting treatment and commercial detergents.展开更多
The alkaline cellulase produced by alkalophilic Bacillus sp. N6-27 was purified to electrophoresis homogeneity by (NH4)2SO4 fractionation, Sepharose CL-4B hydrophobic interaction chromatography, Bio-gel P-150 chromato...The alkaline cellulase produced by alkalophilic Bacillus sp. N6-27 was purified to electrophoresis homogeneity by (NH4)2SO4 fractionation, Sepharose CL-4B hydrophobic interaction chromatography, Bio-gel P-150 chromatography. The molecular weight and pI determined by SDS-PAGE and by PAGE-IEF were 94000 and 4.2,respectively. The optimum temperature and PH for the enzymatic catalysis were 55℃ and 8.5, respectively. The enzyme activity was stable under 50℃ and in the PH range of 6-11. The substrate was carboxymethylcellulose (CMC). The enzyme activity was strongly inhibited by Fe2 +, Cu2+ and Hg2+.展开更多
Chitosanase which catalyze the hydrolytic degradation of chitosan is regarded as an important enzyme for the extense carbon and nitrogen recycle which occurs in nature. A bifunctional enzyme with chitosanase and carbo...Chitosanase which catalyze the hydrolytic degradation of chitosan is regarded as an important enzyme for the extense carbon and nitrogen recycle which occurs in nature. A bifunctional enzyme with chitosanase and carboxymethyl cellulase (CMCase) activity was purified from commercial cellulose, which was produced by Trichoderma viride, through sequential steps of DEAE-Sepharose CL-6B, ion-exchange chromatography, phenyl Sepharose CL-4B hydrophobic chromatography and Sephacryl S-200 gel filtration. The purified hydrolase was homogeneous, as examined by SDS-PAGE and RP-HPLC. The molecular mass was 66 kD and 64 kD estimated by SDS-PAGE and gel filtration,respectively.展开更多
文摘A cellulase producing bacterium (E3 strain) was isolated from fecal matter of elephant and identified as Bacillus sp. using 16S rDNA sequenced based molecular phylogenetic approach. While studying the effect of substrates like Carboxymethyl cellulose (CMC), avicel, starch, maltose, sucrose, glucose, fructose, galactose and lactose on cellulase production, it was found that CMC was best carbon source induced cellulase production followed by lactose in this bacterial strain. A positive synergistic effect of lactose with CMC was also observed with enhancement of 5 - 6 times in cellulase production. The optimum cellulase production was recorded with 1% CMC and 1% lactose when added individually in the Omeliansky’s medium. The results showed that addition lactose with CMC greatly enhances the production and activity of various cellulase enzymes. The optimal fermentation conditions for the biosynthesis of cellulase by this strain were found to be temperature: 37℃, pH 7.0. The nitrogen source NH4Cl at 0.15% was optimum for cellulase production by this bacterium.
基金Qingdao Municipal Science and Technology Commission,Qingdao,China for providing financial support to this work(06-2-2-22-jch)
文摘We have reported that A. pullulans 98 produces a high yield of cellulase. In this study, a carboxymethyl cellulase(CMCase) in the supernatant of the culture of A. pullulans 98 was purified to homogeneity, and the maximum production of CMCase was 4.51 U(mg protein)-1. The SDS-PAGE analysis showed that the molecular mass of the purified CMCase was 67.0 k Da. The optimal temperature of the purified enzyme with considerable thermosensitivity was 40℃, much lower than that of the CMCases from other fungi. The optimal p H of the enzyme was 5.6, and the activity profile was stable in a range of acidity(p H 5.0–6.0). The enzyme was activated by Na+, Mg2+, Ca2+, K+, Fe2+ and Cu2+, however, it was inhibited by Fe3+, Ba2+, Zn2+, Mn2+ and Ag+. Km and Vmax values of the purified enzyme were 4.7 mg m L-1 and 0.57 μmol L-1 min-1(mg protein)-1, respectively. Only oligosaccharides with different sizes were released from carboxymethylcellulose(CMC) after hydrolysis with the purified CMCase. The putative gene encoding CMCase was cloned from A. pullulans 98, which contained an open reading frame of 954 bp(EU978473). The protein deduced contained the conserved domain of cellulase superfamily(glucosyl hydrolase family 5). The N-terminal amino acid sequence of the purified CMCase was M-A-P-H-A-E-P-Q-S-Q-T-T-E-Q-T-S-S-G-Q-F, which was consistent with that deduced from the cloned gene. This suggested that the purified CMCase was indeed encoded by the cloned CMCase gene in this yeast.
文摘Five thermophilic strains that can degrade cellulose were isolated from the compost of a waste management in Guangzhou, China. Since one of them degraded cellulose effectively, it was chosen as the study strain. Based on its morphology, spores′ susceptibility to heat, cell wall composition and other characteristics, the organism was classified as Thermomonospora fusca. Conditions for production of carboxy methyl cellulase (CMCase) were examined. The optimal temperature and pH value for enzyme production were 50 ℃ and 10.5, respectively. Cellulosic materials and easily metabolisable carbohydrates served as carbon sources for the growth of the strain. Only cotton, avicel,carboxy methyl cellulose (CMC) acted as potent inducers for the production of cellulases by this strain. Despite excellent growth on easily metabolisable carbohydrates, only constitutive levels of cellulases were produced. The optimal carbon and nitrogen sources for CMCase production were cotton and soybean respectively. The high thermostability, wide pH stability, and cheap nitrogen source show well potential use for composting treatment and commercial detergents.
文摘The alkaline cellulase produced by alkalophilic Bacillus sp. N6-27 was purified to electrophoresis homogeneity by (NH4)2SO4 fractionation, Sepharose CL-4B hydrophobic interaction chromatography, Bio-gel P-150 chromatography. The molecular weight and pI determined by SDS-PAGE and by PAGE-IEF were 94000 and 4.2,respectively. The optimum temperature and PH for the enzymatic catalysis were 55℃ and 8.5, respectively. The enzyme activity was stable under 50℃ and in the PH range of 6-11. The substrate was carboxymethylcellulose (CMC). The enzyme activity was strongly inhibited by Fe2 +, Cu2+ and Hg2+.
文摘Chitosanase which catalyze the hydrolytic degradation of chitosan is regarded as an important enzyme for the extense carbon and nitrogen recycle which occurs in nature. A bifunctional enzyme with chitosanase and carboxymethyl cellulase (CMCase) activity was purified from commercial cellulose, which was produced by Trichoderma viride, through sequential steps of DEAE-Sepharose CL-6B, ion-exchange chromatography, phenyl Sepharose CL-4B hydrophobic chromatography and Sephacryl S-200 gel filtration. The purified hydrolase was homogeneous, as examined by SDS-PAGE and RP-HPLC. The molecular mass was 66 kD and 64 kD estimated by SDS-PAGE and gel filtration,respectively.
