Figure 6: Analysis of RcsF/IgaA/RcsD interactions.

A) AlphaFold3 model of E.coli RcsF/IgaA/RcsD_1–461 complex. B) Detail of IgaA/RcsD binding interface at the periplasmic end. Interacting residues R506, N509, N513 are shown. C) Detail of contacts between RcsD, RcsF, and IgaA. D) Detail of cytoplasmic contact between IgaA/RcsD. E) AlphaFold3 model of E.coli IgaA/RcsD_1–461 complex, including detailed view of IgaA/RcsD binding interface at the periplasmic end. F) Interaction of IgaA periplasmic mutants with RcsD. The interaction of RcsD was tested with IgaA and IgaA point mutants using BACTH in BTH101. All interactions are plotted relative to RcsD-IgaA interaction (set to 100). G) Signaling by RcsD-binding defective IgaA mutants. The P_rprA:mCherry fluorescence of the IgaA mutant strains was measured to assay the Rcs signaling as described in Fig 3F. RFU at OD 0.4 is plotted and uninduced IgaA is considered as 1. The right panel shows IgaA alleles in the chromosome of a ΔrcsD ΔrcsF strain transformed with pEAW11. H) Signaling by RcsD periplasmic mutants. The ΔrcsD strain (EAW19) was transformed with either pBAD24 vector or RcsD mutants and their fluorescence assayed as described in Fig. 4E.