Abstract
Phytopathogenic Pectobacterium spp. import ferredoxin into the periplasm for proteolytic processing and iron release via the ferredoxin uptake system. Although the ferredoxin receptor FusA and the processing protease FusC have been identified, the mechanistic basis of ferredoxin import is poorly understood. In this work, we demonstrate that protein translocation across the outer membrane is dependent on the TonB-like protein FusB. In contrast to the loss of FusC, loss of FusB or FusA abolishes ferredoxin transport to the periplasm, demonstrating that FusA and FusB work in concert to transport ferredoxin across the outer membrane. In addition to an interaction with the "TonB box" region of FusA, FusB also forms a complex with the ferredoxin substrate, with complex formation required for substrate transport. These data suggest that ferredoxin transport requires energy transduction from the cytoplasmic membrane via FusB both for removal of the FusA plug domain and for substrate translocation through the FusA barrel.IMPORTANCE The ability to acquire iron is key to the ability of bacteria to cause infection. Plant-pathogenic Pectobacterium spp. are able to acquire iron from plants by transporting the iron-containing protein ferredoxin into the cell from proteolytic processing. In this work, we show that the TonB-like protein FusB plays a key role in transporting ferredoxin across the bacterial outer membrane by directly energizing its transport into the cell. The direct interaction of the TonB-like protein with substrate is unprecedented and explains the requirement for the system-specific TonB homologue in the ferredoxin uptake system. Since multiple genes encoding TonB-like proteins are commonly found in the genomes of Gram-negative bacteria, this may be a common mechanism for the uptake of atypical substrates via TonB-dependent receptors.