Fig. 1: The cysteine residues in the periplasmic C-tail of SprF are important for SprB movement and gliding motility.
From: A shared mechanism for Bacteroidota protein transport and gliding motility
a Current model for the organization of the gliding motility apparatus and T9SS in F. johnsoniae involving a shared Energy Chain. OM, outer membrane. IM, inner membrane. b Sequence of the periplasmic C-tail of F. johnsoniae SprF with the twin-cysteine motif highlighted and a Weblogo display of the sequence conservation (500 closest whole protein homologs) above. c–e Characterization of sprF mutants with substitutions in the conserved cysteine pair. wt, wild type. Similar results were obtained for 3 biological repeats. c Spreading (gliding) morphology on agar. Scale bar, 5 mm. d Cell surface exposure of the adhesin SprB assessed by protease protection. The indicated strains expressing a fusion between HaloTag and the final 448 amino acids of SprB (Halo-SprB448) were incubated with Proteinase K ( + lanes). Reactions were stopped immediately (t0) or after 20 min (t20) and analysed by immunoblotting with Halotag antibodies. The cytoplasmic protein GroEL serves as both a loading and cell integrity control. In each panel (which correspond to separate immunoblots) a lane has been removed between the t0 and t20 samples. e Exemplar trajectories of single fluorophore-labeled HaloTag-SprB adhesin molecules or HaloTag-SprC molecules in the indicated backgrounds. The trajectories link the single molecule localizations in successive frames recorded for agarose-immobilized cells and are 2D projections of what is a 3D process. Each trajectory within a cell is shown in a different color and the segmented outline of the cell is shown in black. For the SprF cysteine variants cells were chosen to illustrate the observed range of SprB mobility behaviors. See also Supplementary Movie 1.
