Novel features of the ISC machinery revealed by characterization of Escherichia coli mutants that survive without iron–sulfur clusters

Summary Biological assembly of iron–sulfur (Fe–S) clusters is mediated by complex systems consisting of multiple proteins. Escherichia coli possesses two distinct systems called the ISC and SUF machineries encoded by iscSUA‐hscBA‐fdx‐iscX and sufABCDSE respectively. Deletion of both pathways results in absence of the biosynthetic apparatus for Fe–S clusters, and consequent lethality, which has hampered detailed genetic studies. Here we report that modification of the isoprenoid biosynthetic pathway can offset the indispensability of the Fe–S cluster biosynthetic systems and show that the resulting Δisc Δsuf double mutants can grow without detectable Fe–S cluster‐containing proteins. We also constructed a series of mutants in which each isc gene was disrupted in the deletion background of sufABCDSE. Phenotypic analysis of the mutants revealed that Fdx, an essential electron‐transfer Fe–S protein in the ISC machinery, is dispensable under anaerobic conditions, which is similar to the situation with IscA. Furthermore, we found that several suppressor mutations in IscU, an Fe–S scaffold protein responsible for the de novo Fe–S cluster assembly, could bypass the essential role of the chaperone system HscA and HscB. These findings pave the way toward a detailed molecular analysis to understand the mechanisms involved in Fe–S cluster biosynthesis. We report here that biosynthesis of iron‐sulfur (Fe‐S) cluster is not an essential task for Escherichia coli mutant in which isoprenoid biosynthetic pathway was modified. Genetic characterization of the ISC components in a SUF‐deficient background revealed that Fdx is dispensable for the Fe‐S cluster assembly under anaerobic conditions and that the essential role of HscA/HscB chaperone system can be bypassed by several point mutations in IscU.