Mutational and structural studies of the active-site residues in truncated Fibrobacter succinogenes1,3-1,4-beta-D-glucanase.

Tsai LC; Huang HC; Hsiao CHChiang YNShyur LFLin YSLee SH

首页> 外文期刊>Acta crystallographica.Section D. Biological crystallography >Mutational and structural studies of the active-site residues in truncated Fibrobacter succinogenes1,3-1,4-beta-D-glucanase.

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Mutational and structural studies of the active-site residues in truncated Fibrobacter succinogenes1,3-1,4-beta-D-glucanase.

机译：突变和结构的研究在截断Fibrobacter活性位点残基3 - 1, succinogenes1 4-beta-D-glucanase。

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1,3-1,4-beta-D-Glucanases (EC 3.2.1.73) specifically hydrolyze beta-1,4-glycosidic bonds located prior to beta-1,3-glycosidic linkages in lichenan or beta-D-glucans. It has been suggested that truncated Fibrobacter succinogenes 1,3-1,4-beta-D-glucanase (TFsbeta-glucanase) can accommodate five glucose rings in its active site upon enzyme-substrate interaction. In this study, 12 mutant enzymes were created by mutating the conserved residues Gln70, Asn72, Gln81 and Glu85 proposed to bind to substrate subsites +1 and +2 and the catalytic properties of these mutants were determined. The most significant change in catalytic activity was observed on mutation of Gln70, with a 299-fold and 498-fold lower k(cat)/K(m) for the mutants Q70A and Q70I, respectively, compared with the wild-type enzyme. Mutagenesis, kinetic and structural studies revealed that the conserved residues surrounding the active site of TFsbeta-glucanase at substrate subsites +1 and +2 play an important role in its catalytic function, with the following order of importance: Gln70 > Asn72 > Glu85 > Gln81. The crystal structure of mutant E85I was determined at 2.2 A resolution. Further analysis of the E85I mutant structure revealed that the loop located at the concave site moved approximately 2 A from its position in the native enzyme complex without changing the core structure.

机译：1、3 - 1、4-beta-D-Glucanases (EC 3.2.1.73)特别是水解beta 1, 4-glycosidic债券坐落在beta 1之前,3-glycosidic联系lichenan或beta-D-glucans。,截断Fibrobacter succinogenes1、3 - 1、4-beta-D-glucanase (TFsbeta-glucanase)容纳5葡萄糖环的活性部位在es交互。12突变酶是由突变守恒的残留Gln70、Asn72 Gln81 Glu85提出结合基质子+ 1和+ 2这些突变体的催化性能测定。观察催化活性的突变Gln70,低299倍和498倍k(猫)/ k (m)的突变体Q70A Q70I,与野生型相比,酶。诱变、动力学和结构研究发现周围的守恒的残留物TFsbeta-glucanase在底物的活性部位子站+ 1和+ 2起着重要的作用催化功能,以下的顺序重要性:Gln70 > Asn72 > Glu85 > Gln81。晶体结构的变异E85I决心2.2一项决议。突变体结构显示,循环凹部位移动大约2它的位置在本机酶复杂改变的核心结构。

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《Acta crystallographica.Section D. Biological crystallography》 |2008年第12期|1259-1266|共8页
作者
Tsai LC; Huang HC; Hsiao CHChiang YNShyur LFLin YSLee SH;
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Department of Molecular Science and Engineering, National Taipei University of Technology, Taipei 106, Taiwan. lichu@ntut.edu.tw;

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正文语种英语
中图分类化学;
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lichenin; Substrate Interaction; Mutantscore structureFibrobacterSTRUCTURAL STUDYenzyme complexesFibrobacter succinogenesActive Site;

机译：苔粉;基质交互;MutantscorestructureFibrobacterSTRUCTURAL STUDYenzymecomplexesFibrobacter succinogenesActive网站;

Mutational and structural studies of the active-site residues in truncated Fibrobacter succinogenes1,3-1,4-beta-D-glucanase.

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