首页> 外文期刊>Antonie van Leeuwenhoek: Journal of Microbiology and serology >An acidothermophilic functionally active novel GH12 family endoglucanase from Aspergillus niger HO: purification, characterization and molecular interaction studies
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An acidothermophilic functionally active novel GH12 family endoglucanase from Aspergillus niger HO: purification, characterization and molecular interaction studies

机译:来自Aspergillus Niger Ho的酸热功能活性新的GH12家族内葡聚糖酶:纯化,表征和分子相互作用研究

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摘要

Endoglucanase (EG) from Aspergillus niger HO was sequentially purified through ultrafiltration, ion exchange and size exclusion chromatography to homogeneity, with an overall recovery of 18 %. The purified EG was a monomeric protein with a molecular weight of about 55 kDa. The enzyme was optimally active at pH 3.5 and 70 A degrees C with a half life (t(1/2)) of 3 h and K-m value of 2.5 mg/ml. Metal ions, such as Ca2+ and Co2+ helped in enzyme induction, while Hg2+ and Cu2+ strongly inhibited the enzyme activity. Peptide mass fingerprinting results revealed that the purified EG is a novel enzyme that belongs to family 12 of glycoside hydrolase (GH12). Molecular docking studies indicated the presence of Glu116 and Glu204 as important determinant residues for the functional interaction with carboxymethylcellulose and showed hydrogen bonding with Asp99, Glu116, Glu204 and hydrophobic interactions with Trp22, Val58, Tyr61, Phe101, Met118, Trp120, Pro129, Ile130, Thr160 and Phe206. Hydrolysis of 2 % CMC with purified acidothermophilic EG at its optimum temperature and pH resulted in complete hydrolysis within 2 h yielding 18 % cellotriose, 72 % cellobiose and 10 % glucose as evident from HPLC analysis. In comparison to most of the EGs reported in literature, EG from A. niger HO exhibited higher thermostability. The acidothermophilic nature of this enzyme makes it potentially useful for industrial applications.
机译:通过超滤,离子交换和尺寸排阻色谱法向均匀性依次纯化来自曲霉血清葡萄糖酶(例如),总体回收率为18%。纯化的例如分子量为约55kDa的单体蛋白质。酶在pH 3.5和70℃下最佳活性活性,半衰期(T(1/2))为3小时和K-M值为2.5mg / mL。金属离子,例如Ca2 +和CO 2 +有助于酶诱导,而Hg2 +和Cu2 +强烈抑制酶活性。肽质量指纹识别结果表明,纯化的例如属于糖苷水解酶(GH12)的家庭12的新酶。分子对接研究表明了Glu116和Glu204的存在作为与羧甲基纤维素的功能相互作用的重要决定性残基,并与ASP99,GLU116,GLU204和与TRP22,VAL58,TYR61,PHE101,MET118,TRP120,PRO129,ILE130,PRO129,ILE130的疏水相互作用显示氢键。 thr160和phe206。将2%CMC的水解具有纯化的酸热剂,例如在其最佳温度和pH下导致2小时内的完全水解,从HPLC分析中明显表示18%Cellotriose,72%的纤维二糖和10%葡萄糖。与文献中报告的大多数EGS相比,例如来自A.尼日尔浩的热稳定性。该酶的酸热性质使其可能适用于工业应用。

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