SmsB-SmsC machinery in Thermococcus kodakarensis functions as an archaeal scaffold mediating Fe-S cluster assembly

The SUF system is one of the multiprotein machineries responsible for iron-sulfur (Fe-S) cluster biogenesis. In bacterial and eukaryotic SUF systems, SufB, SufC, and SufD form a SufBC₂D complex as the scaffold for Fe-S cluster assembly. SMS (SUF-like minimal system), composed only of SufB and SufC homologs (SmsB and SmsC) without a SufD component, has recently been established, and representatives from methanogenic archaea have been experimentally verified to function in Fe-S cluster biogenesis. SMS has been proposed to be an ancestor of the SUF system. The hyperthermophilic archaeon Thermococcus kodakarensis harbors candidate proteins for SmsB and SmsC (Tk-SmsB and Tk-SmsC) encoded by TK0730 and TK0731 genes, respectively. As Tk-SmsB is phylogenetically positioned in a clade distinct from the previously characterized SmsB proteins from methanogens, here, we examined whether Tk-SmsB and Tk-SmsC can also function in Fe-S cluster generation and transfer. Tk-SmsB and Tk-SmsC formed a heterotetrameric SmsB₂C₂ complex. Tk-SmsC displayed ATPase activity, and its catalytic efficiency (k_cat/K_m) increased up to 10-fold upon complex formation with Tk-SmsB. The Tk-SmsB₂C₂ complex could generate Fe-S clusters in the presence of ferric and sulfide ions. The holo-Tk-SmsB₂C₂ complex could transfer the Fe-S clusters to the apo-form of the Fe-S cluster-dependent lipoyl synthase LipS from T. kodakarensis, resulting in a LipS with sulfur insertion activity for lipoyl group biosynthesis. In the presence of cysteine desulfurase and ATP, the apo-Tk-SmsB₂C₂ complex could also generate Fe-S clusters utilizing cysteine as a sulfur donor and activate LipS.IMPORTANCEA representative of an SmsB protein from T. kodakarensis (Tk-SmsB) that lies in a clade phylogenetically distinct from those of previously verified SmsB proteins has been examined. The results demonstrate that Tk-SmsB, along with Tk-SmsC, functions as a scaffold for Fe-S cluster synthesis in T. kodakarensis, adding further support to the proposition that SMS represents the primitive form of the SUF systems widely present in bacteria and eukaryotes.