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嗜热菌Thermus thermophilus HB8铁硫簇合成途径中IscA和IscU蛋白的表达、纯化和功能研究

Fuctions and Reconstitution of IscA and IscU in the Fe-S Cluster Biosynthesis Pathway from the Thermus thermophilus HB8
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摘要 Fe-S簇可参与电子传递和基因调节等过程.实验以嗜热菌Thermus thermophilus HB8基因组为模板,通过PCR扩增获得Fe-S簇ISC系统生物合成途径中的IscA和IscU基因,并克隆连接到表达载体pET-28a(+)上.重组质粒转化E.coli Rosetta(DE3)进行表达,用亲和层析纯化得到浓度和纯度都较高的IscA和IscU蛋白.SDS-PAGE分析,证实其分子量大小分别为13.7KD和14.8KD.实验确定IscA是单聚体,IscA上能形成铁硫簇,IscU上不能形成铁硫簇.在厌氧条件下,RNA的量能提高1.5倍,但是IscA和IscU的表达量都会降低,说明厌氧对这两个基因有损伤. Iron-sulfur cluster plays an important role in electron transfer and gene regulation. In this study,the gene of IscA and IscU from HB8 were cloned and expressed in Rosetta,and the proteins were purified by using affinity chromatography. SDS-PAGE revealed the moleculer weight of IscA was 13. 7KD and IscU was 14. 8KD. Iron sulfur cluster can be formed on IscA,Iron sulfur cluster can＇t be formed on IscU,under anaerobic conditions,RNA can increase 1. 5 times,But the amount of expression of IscA and IscU will be reduced,indicating that the anaerobic damage on these two gene.
作者 吴琦 谭菲 刘晓晴
机构地区 首都师范大学生命科学学院
出处 《首都师范大学学报（自然科学版）》 2014年第2期49-56,共8页 Journal of Capital Normal University：Natural Science Edition
关键词 嗜热菌Thermus THERMOPHILUS HB8 克隆 体外重组. Thermus thermophilus HB8 cloning reconstitution in vitro.
分类号 Q78 [生物学—分子生物学]
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参考文献16

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首都师范大学学报（自然科学版）
2014年 第2期

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