Receptor-substrate competition for the TonB homolog FusB suggests a model for ferredoxin import

TY - JOUR

T1 - Receptor-substrate competition for the TonB homolog FusB suggests a model for ferredoxin import

AU - Wojnowska, Marta

AU - Flores, Victor

AU - Yelland, Tamas

AU - Fisher, Stuart R.

AU - Bogucka, Agnieszka

AU - Stott, Katherine

AU - Walker, Daniel

N1 - Copyright: © 2026 The Authors. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology. This is an open access article under the CC BY license. http://creativecommons.org/licenses/by/4.0/

PY - 2026/6

Y1 - 2026/6

N2 - TonB-dependent uptake systems of gram-negative bacterial pathogens constitute prominent virulence factors, allowing nutrients—primarily siderophore-bound iron—to cross the highly impermeable outer membrane. Remarkably, the ferredoxin uptake system (Fus) of certain soft rot plant pathogens imports an entire host protein into the periplasm and extracts its bound iron for growth. The inner membrane protein FusB, a TonB homolog, plays two roles in facilitating import. First, like other TonBs, it remodels the globular plug domain obstructing the lumen of the outer membrane receptor FusA to allow ferredoxin passage. Unusually for a TonB protein, FusB then interacts directly with the FusA-bound ferredoxin substrate to facilitate its transport into the periplasm. Here, we describe structures of the FusB homodimer as well as the FusB-ferredoxin complex and, using biophysical, biochemical, and mutagenesis approaches, we determine the key features of the binding interfaces formed by FusB with FusA and ferredoxin. The C-terminal domain of FusB (FusB-CTD) exists in a monomer-dimer equilibrium in vitro, with the homodimer stabilized by an intermolecular R241-D322 salt bridge. The “FusB-box” of FusA interacts with monomeric FusB-CTD, and FusA D53 outcompetes FusB D322 to bind R241. Upon ferredoxin binding, FusB-CTD undergoes a structural rearrangement, expanding its β-sheet from three to four strands. In agreement with the proposed sequence of events, ferredoxin binding displaces FusA from FusB with R241 forming an intramolecular salt bridge with D322 to stabilize the newly formed β-hairpin of FusB. We propose a mechanistic model for ferredoxin import where FusB R241 acts as a molecular switch.

AB - TonB-dependent uptake systems of gram-negative bacterial pathogens constitute prominent virulence factors, allowing nutrients—primarily siderophore-bound iron—to cross the highly impermeable outer membrane. Remarkably, the ferredoxin uptake system (Fus) of certain soft rot plant pathogens imports an entire host protein into the periplasm and extracts its bound iron for growth. The inner membrane protein FusB, a TonB homolog, plays two roles in facilitating import. First, like other TonBs, it remodels the globular plug domain obstructing the lumen of the outer membrane receptor FusA to allow ferredoxin passage. Unusually for a TonB protein, FusB then interacts directly with the FusA-bound ferredoxin substrate to facilitate its transport into the periplasm. Here, we describe structures of the FusB homodimer as well as the FusB-ferredoxin complex and, using biophysical, biochemical, and mutagenesis approaches, we determine the key features of the binding interfaces formed by FusB with FusA and ferredoxin. The C-terminal domain of FusB (FusB-CTD) exists in a monomer-dimer equilibrium in vitro, with the homodimer stabilized by an intermolecular R241-D322 salt bridge. The “FusB-box” of FusA interacts with monomeric FusB-CTD, and FusA D53 outcompetes FusB D322 to bind R241. Upon ferredoxin binding, FusB-CTD undergoes a structural rearrangement, expanding its β-sheet from three to four strands. In agreement with the proposed sequence of events, ferredoxin binding displaces FusA from FusB with R241 forming an intramolecular salt bridge with D322 to stabilize the newly formed β-hairpin of FusB. We propose a mechanistic model for ferredoxin import where FusB R241 acts as a molecular switch.

KW - bacterial pathogenesis

KW - biophysics

KW - gram-negative bacteria

KW - membrane transport

KW - molecular biology

KW - protein complex

KW - protein crystallization

KW - protein translocation

UR - https://www.scopus.com/pages/publications/105040721746

U2 - 10.1016/j.jbc.2026.113121

DO - 10.1016/j.jbc.2026.113121

M3 - Article

C2 - 42103222

AN - SCOPUS:105040721746

SN - 0021-9258

VL - 302

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 6

M1 - 113121

ER -