Mechanism of iron-sulfur cluster assembly : Structure and functional interaction of Isc proteins

点击翻译按钮获取中文摘要

Iron-sulfur proteins are present in almost all living organisms and exhibit diverse functions, which include electron transport, redox and non-redox catalysis, and sensing for regulatory processes. They contain clusters of iron and sulfur atoms with vailiable complexity, such as [2Fe-2S], [3Fe-4S], and [4Fe-4S] clusters. Still very little is known about the in vivo synthesis of these clusters, which is believed to require a complex machinery encoded us prokaryotes by the isc (iron suffer cluster) operon, iscRSUA-hscBA-fdx-ORF3. We have undertaken genetic and biochemical approaches to elucidate the complex mechanism by which Fe-S dusters are assembled.1.Essential compornents of the ISC machinery (IscS, IscU, HscB, HscA, and Fdx) have been identified by a systematic mutational analysis of the isc operon in Eseherichia coli.2.Biochemical characterization of the recombinant IscA led to the identification of labile [2Fe-2S] cluster and specific interaction with Fdx.3.We determined the crystal structure of Fdx, which suggested the important role of Cys46 located on the molecular susface in the vicinity of [2Fe-2SJ cluster.4.Network of protein-protein interactions among the components of the ISC machinery has been elucidated by the combination of two-hybrid and pull-down analyses.5.We have isolated a covalently-bound IscS/IscU complex and identified that Cys328 of IseS and Cys63 of lscU are the sites of disulfide bridge formation.6.We have analyzed spontaneous psuedorevertants isolated from a mutant shale lacking all the components of the ISC machinery, which led to the identification of the sufABCDSE (suf) operon participating in the ISC-independent pathway for the assembly of Fe-S clusters. The genes homologous to sufBC are present in a wide range of bacteria, archaea and plastids, suggesting that this type of system is almost ubiquitous in nature.