嗜酸氧化亚铁硫杆菌铁硫簇组装蛋白IscU的表达、纯化及特性分析

Expression, purification and characterization of an iron-sulfur cluster assembly protein, IscU, from Acidithiobacillus ferrooxidans.

作者信息

Zeng Jia, Zhao Wenjie, Liu Yuandong, Xia Lexian, Liu Jianshe, Qiu Guanzhou

机构信息

Department of Bioengineering, School of Resources Processing and Bioengineering, Central South University, Changsha, 410083, PR China.

出版信息

Biotechnol Lett. 2007 Dec;29(12):1965-72. doi: 10.1007/s10529-007-9488-1. Epub 2007 Jul 28.

DOI:10.1007/s10529-007-9488-1

PMID:17660945

Abstract

An iron-sulfur cluster assembly protein, IscU, is encoded by the operon iscSUA in Acidithiobacillus ferrooxidans. The gene of IscU was cloned and expressed in Escherichia coli. The protein was purified by one-step affinity chromatography to homogeneity. The protein was in apo-form, the [Fe(2)S(2)] cluster could be assembled in apoIscU with Fe(2+) and sulfide in vitro, and in the presence of IscA and IscS, the IscU could utilize L: -cysteine and Fe(2+) to synthesize [Fe(2)S(2)] cluster in the protein. Site-directed mutagenesis for the protein revealed that Cys37, Asp39, Cys63 and Cys106 were involved in ligating with the [Fe(2)S(2)] cluster.

摘要

铁硫簇组装蛋白IscU由嗜酸氧化亚铁硫杆菌中的iscSUA操纵子编码。IscU基因在大肠杆菌中克隆并表达。该蛋白通过一步亲和层析纯化至均一性。该蛋白为脱辅基形式，[Fe(2)S(2)]簇可在体外与Fe(2+)和硫化物在脱辅基IscU中组装，并且在IscA和IscS存在的情况下，IscU可利用L-半胱氨酸和Fe(2+)在蛋白中合成[Fe(2)S(2)]簇。对该蛋白的定点诱变表明，Cys37、Asp39、Cys63和Cys106参与与[Fe(2)S(2)]簇的连接。