The ErpA/NfuA complex builds an oxidation-resistant Fe-S cluster delivery pathway

Fe-S cluster-containing proteins occur in most organisms, wherein they assist in myriad processes from metabolism to DNA repair via gene expression and bioenergetic processes. Here, we used both in vitro and in vivo methods to investigate the capacity of the four Fe-S carriers, NfuA, SufA, ErpA, and IscA, to fulfill their targeting role under oxidative stress. Likewise, Fe-S clusters exhibited varying half-lives, depending on the carriers they were bound to; an NfuA-bound Fe-S cluster was more stable (t(1/2) = 100 min) than those bound to SufA (t(1/2) = 55 min), ErpA (t(1/2) = 54 min), or IscA (t(1/2) = 45 min). Surprisingly, the presence of NfuA further enhanced stability of the ErpA-bound cluster to t(1/2) = 90 min. Using genetic and plasmon surface resonance analyses, we showed that NfuA and ErpA interacted directly with client proteins, whereas IscA or SufA did not. Moreover, NfuA and ErpA interacted with one another. Given all of these observations, we propose an architecture of the Fe-S delivery network in which ErpA is the last factor that delivers cluster directly to most if not all client proteins. NfuA is proposed to assist ErpA under severely unfavorable conditions. A comparison with the strategy employed in yeast and eukaryotes is discussed.