Genetic analysis of the isc operon in Escherichia coli involved in the biogenesis of cellular iron-sulfur protein
Genetic analysis of the isc operon in Escherichia coli involved in the biogenesis of cellular iron-sulfur protein
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DOI：
10.1093/oxfordjournals.jbchem.a002963
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发表时间：
2001-07-01
影响因子：
2.7
通讯作者：
Takahashi, Y
中科院分区：
文献类型：
--
作者：
Tokumoto, U;Takahashi, Y
铁硫（Fe-S）簇是许多蛋白质活性所必需的非血红素铁辅因子，它在复杂机制的帮助下被整合到靶蛋白中。在细菌中，已经提出由iscRSUA-hscBA-fdx-ORF 3簇（isc操纵子）编码的几种蛋白质在Fe-S簇的组装中执行关键任务。为了阐明体内功能，我们对大肠杆菌伊势操纵子中的基因进行了系统的突变分析。在所有功能测试中，即生长速率、营养需求和Fe-S酶活性，iscS基因的失活引起了最剧烈的改变。在iscU，hscB，hscA和fdx基因突变的菌株也表现出明显的表型结果几乎彼此相同。iscA的失活对Fe-S酶的影响很小，但明显。相反，具有失活的iscR或ORF 3的突变体与野生型细胞几乎没有差异。通过使用缺失整个伊势操纵子的突变株的互补实验进一步证实了iscSUA-hscBA-fdx用于FeS簇组装的要求。我们的研究结果支持这样的结论，即IscS，通过半胱氨酸脱硫酶活性，提供硫，该硫随后通过由iscUA-hscBA-fdx基因产物组成的组装机器并入Fe-S簇中。这里呈现的结果表明IscU，HscB，HscA和Fdx作为组装机器的中心组分的关键作用，并且还为它们之间的相互作用提供证据。
The iron-sulfur (Fe-S) cluster, the nonheme-iron cofactor essential for the activity of many proteins, is incorporated into target proteins with the aid of complex machinery. In bacteria, several proteins encoded by the iscRSUA-hscBA-fdx-ORF3 cluster (isc operon) have been proposed to execute crucial tasks in the assembly of Fe-S clusters, To elucidate the in vivo function, we have undertaken a systematic mutational analysis of the genes in the Escherichia coli ise operon. In all functional tests, i,e, growth rate, nutritional requirements and activities of Fe-S enzymes, the inactivation of the iscS gene elicited the most drastic alteration. Strains with mutations in the iscU, hscB, hscA, and fdx genes also exhibited conspicuous phenotypical consequences almost identical to one another. The effect of the inactivation of iscA was small hut appreciable on Fe-S enzymes, In contrast, mutants with inactivated iscR or ORF3 showed virtually no differences from wild-type cells. The requirement of iscSUA-hscBA-fdx for the assembly of FeS clusters was further confirmed by complementation experiments using a mutant strain in which the entire ise operon was deleted. Our findings support the conclusion that IscS, via cysteine desulfurase activity, provides the sulfur that is subsequently incorporated into Fe-S clusters by assembler machinery comprising of the iscUA-hscBA-fdx gene products, The results presented here indicate crucial roles for IscU, HscB, HscA, and Fdx as central components of the assembler machinery and also provide evidence for interactions among them.