Fig. 4: LpoB preferentially promotes glycan synthesis initiation. | Nature Communications

Fig. 4: LpoB preferentially promotes glycan synthesis initiation.

From: The hit-and-run of cell wall synthesis: LpoB transiently binds and activates PBP1b through a conserved allosteric switch

Fig. 4

a Schematic illustrating the PBP1b-LpoB smFRET binding assay. Cy3-labeled PBP1b is tethered to the surface, whereas Cy5-labeled LpoB is supplied in solution. In this set-up, Cy5 fluorescence and FRET signal are detected only when LpoB binds PBP1b. b Example trajectories, showing donor and acceptor fluorescence upon donor excitation (top), acceptor fluorescence upon acceptor excitation (middle) and the calculated FRET efficiency signal (bottom). Anti-correlated changes in donor and acceptor fluorescence (top) correspond to FRET transitions (bottom) and perfectly correlate with changes in direct acceptor signal (middle), i.e., Cy5-Cy3 colocalization events. c Example trajectories of LpoB binding either to apo PBP1b or to PBP1b complexed with lipid II substrate or glycan chains. Step-like increases and decreases in FRET efficiency correspond to association and dissociation events, respectively. d Dwell time histograms and exponential fits of samples from (c) with mean fits indicated on the figure. Insets show enlarged views of the histograms and fits in the 0 to 5 s range. Full binding statistics are summarized in the SI Table 3. e Schematic illustrating the relative changes in the stability of the PBP1b-LpoB complex throughout the polymerization reaction.

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