PilB 12-163 -PilZ and PilB 1-190 -PilZ can form ternary complexes with FimX EAL and FimX GGDEF-EAL fragments.

Size exclusion chromatography (Superdex 200 resin, 10/300 column) analysis of the interactions of PilB 12-163 -PilZ and PilB 1-190 -PilZ complexes with different FimX fragments. Chromatograms are shown for the PilB 12-163 -PilZ or PilB 1-190 -PilZ complexes on their own (blue), FimX fragments on their own (green) and 1:1 mixtures PilB-PilZ and FimX fragments (100 μM) in the absence (red) or presence (black) of c-di-GMP (2-fold excess of c-di-GMP to FimX). ( A ) PilB 12-163 -PilZ complex and FimX EAL . ( B ) PilB 1-190 -PilZ complex and FimX EAL . ( C ) PilB 12-163 -PilZ complex and FimX GGDEF-EAL . ( D ) PilB 1-190 -PilZ complex and FimX GGDEF-EAL . ( E ) PilB 12-163 -PilZ complex and FimX PAS-GGDEF-EAL . ( F ) PilB 12-163 -PilZ complex and full-length FimX. SDS-PAGE analysis of representative fractions is shown on the right of each panel (vertical dotted lines in D, E and F indicate parts of the gel removed between molecular mass markers and protein samples). Ternary complexes are observed when using FimX EAL and FimX GGDEF-EAL but not when using FimX PAS-GGDEF-EAL or full-length FimX. Each experiment was performed at least three times and representative results are shown. (TIF)