BIOPROCESS AND BIOSYSTEMS ENGINEERING
·
2020
·Vol. 44
·期 1
·67-79
·
IF 2.419
Zhou Hai-Yan,Zhou Jian-Bao,Yi Xiao-Nan,Wang Yan-Mei,Xue Ya-Ping,Chen De-Shui,Cheng Xin-Ping,Li Mian,Wang Hong-Yan,Chen Kai-Qian,Liu Zhi-Qiang,Zheng Yu-Guo
To develop new cellulases for efficient utilization of the lignocellulose, an endoglucanase (CoCel5A) gene from Colletotrichum orchidophilum was synthesized and a recombinant Pichia pastoris GS115/pPIC9K/cocel5A was constructed for secretory expression of CoCel5A. After purification, the protein CoCel5A was biochemically characterized. The endoglucanase CoCel5A exhibited the optimal activity at 55–75 °C and high thermostability (about 85% residual activity) at the temperature of 55 °C after incubation for 3 h. The highest activity of CoCel5A was detected when 100 mM citric acid buffer (pH 4.0–5.0) was used and excellent pH stability (up to 95% residual activity) was observed after incubation in 100 mM citric acid buffer (pH 3.0–6.0) at 4 °C for 24 h. Carboxymethyl cellulose sodium salt ( n = approx. 500) (CMC) and β - d -glucan were the best substrates for CoCel5A among the tested substrates. The kinetic parameters V max , K m , and K cat / K m values against CMC were 290.70 U/mg, 2.65 mg/mL, and 75.67 mL/mg/s, respectively; and 228.31 U/mg, 2.06 mg/mL, and 76.45 mL/mg/s against β - d -glucan, respectively, suggesting that CoCel5A has high affinity and catalytic efficiency. These properties supported the potential application of CoCel5A in biotechnological and environmental fields.
研究领域:
生物化学
·
生物技术
·
酶学
·
微生物学
✓ 已验证
DOI
PubMed
高影响因子
TEXTILE RESEARCH JOURNAL
·
2017
·
IF 1.443
Ruijun Wang,Zhengchu Liu,Lifeng Cheng,Shengwen Duan,Xiangyuan Feng,Ke Zheng,Yi Cheng,Jie Zeng
Xylanase plays an important role in the hydrolysis of hemicellulose and has gained much attention in the field of biological degumming. The research for xylanases with cellulase-free and high activity for biological degumming has intensified in recent years. In the present research, heterologous expression of a novel endo-β-1,4-xylanase (GH30) from Dickeya dadantii DCE-01 in Escherichia coli BL21 (DE3) was reported. Biochemical characterization of the enzyme and a potential application in ramie biological degumming was discussed. The results showed that the xylanase gene consists of 1251 nucleotides, belonging to glycoside hydrolase family 30 (GH30). The optimal activity of the xylanase was observed at 50℃ and a pH value of 6.4. The Km and Vmax values for beechwood xylan were 14.25 mg/mL and 296.6 μmol/mg, respectively. The catalytic activity was enhanced by addition of 1 mM Cu2+, Ca2+, Mg2+, and K+. The recombinant enzyme was specific for xylan substrates. The enzyme exhibited hydrolytic activity toward ramie hemicellulose. The recombinant xylanase could be effectively applied to ramie degumming.
研究领域:
生物化学
·
生物技术
·
酶学
·
纺织科学
✓ 已验证
DOI
高影响因子
APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY
·
2022
·Vol. 194
·期 12
·6021-6036
·
IF 3.094
Zhang Xin,Wang Yibing,Liu Jiaqi,Wang Weiyang,Yan Xuecui,Zhou Yifa,Cui Jing,Yuan Ye
Because β-1,6-galactans are significant components in arabinogalactans from plant cell walls, identifying selective endo-β-1,6-galactanases is crucial to degrading these polysaccharides and to analyzing and modifying their structures. Here, we cloned and expressed in E. coli a novel endo-β-1,6-galactanase in the glycosidic hydrolase family 30 (GH30) from Penicillium oxalicum . Our recombinant PoGal30 hydrolase (1464 bp gene) that contains an N-terminal His-tag for purification by nickel affinity chromatography has a specific activity of 3.8 U/mg on the substrate de-arabinosylated gum Arabic (dGA) polysaccharide. The enzyme has 487 residues with a molecular mass of 60 kDa, an isoelectric point of 6, and functional pH and temperature optima of pH 2.5 to pH 5.0 and 40 °C, respectively. While the activity of PoGal30 is activated by Mg 2+ (5 or 50 mmol/L), it is completely inhibited by Cu 2+ and Fe 3+ (50 mmol/L) and partially inhibited by Hg 2+ , EDTA, and SDS (50 mmol/L). The enzyme demonstrates high specificity towards β-1,6-galactosidic linkages in dGA, but is inactive against aryl-glycosides and galactobioses with different linkages. Using PoGal30 is, therefore, an effective approach to analyzing the fine structure of polysaccharides and preparing bioactive oligosaccharides.
研究领域:
生物化学
·
分子生物学
·
酶学
·
植物细胞壁分析
✓ 已验证
产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）