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青春双歧杆菌木糖异构酶的表达、纯化、结晶及初步X射线衍射分析

Expression, purification, crystallization and preliminary X-ray diffraction analysis of Bifidobacterium adolescentis xylose isomerase.

作者信息

Dos Reis Caio Vinicius, Bernardes Amanda, Polikarpov Igor

机构信息

Instituto de Física de São Carlos, Universidade de São Paulo, Trabalhador São Carlense 400, 13566-590 São Carlos-SP, Brazil.

出版信息

Acta Crystallogr Sect F Struct Biol Cryst Commun. 2013 May 1;69(Pt 5):588-91. doi: 10.1107/S174430911301110X. Epub 2013 Apr 30.

Abstract

Xylose isomerase (EC 5.3.1.5) is a key enzyme in xylose metabolism which is industrially important for the transformation of glucose and xylose into fructose and xylulose, respectively. The Bifidobacterium adolescentis xylA gene (NC_008618.1) encoding xylose isomerase (XI) was cloned and the enzyme was overexpressed in Escherichia coli. Purified recombinant XI was crystallized using the sitting-drop vapour-diffusion method with polyethylene glycol 3350 as the precipitating agent. A complete native data set was collected to 1.7 Å resolution using a synchrotron-radiation source. The crystals belonged to the orthorhombic space group P21212, with unit-cell parameters a = 88.78, b = 123.98, c = 78.63 Å.

摘要

木糖异构酶(EC 5.3.1.5)是木糖代谢中的关键酶,在工业上分别将葡萄糖和木糖转化为果糖和木酮糖具有重要意义。克隆了编码木糖异构酶(XI)的青春双歧杆菌木糖A基因(NC_008618.1),并在大肠杆菌中对该酶进行了过量表达。使用聚乙二醇3350作为沉淀剂,通过坐滴气相扩散法对纯化的重组XI进行结晶。使用同步辐射源收集了完整的天然数据集,分辨率达到1.7 Å。晶体属于正交空间群P21212,晶胞参数a = 88.78,b = 123.98,c = 78.63 Å。

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