Abstract
The MukBEF protein complex in Escherichia coli is a condensin-like system that binds to bacterial chromosomal DNA. Using bespoke slimfield millisecond fluorescence imaging we were able to visualize the three MukB, E and F protein components in functional complexes in single live cells, both separately as genomic YPet fusions and multiply as dual-label GFP/mCherry mutants and obtain in vivo estimates for the stoichiometry of these components using step-wise photobleaching of fluorescent proteins, precise down to the level of single molecules. This indicates a predominately tetrameric complex of B:E:F = 4:4:2 suggesting that the DNA-bound MukBEF in the live cell is in its ATP-bound state, and which is highly dynamic in the living cell with focal complexes undergoing molecular turnover over a time scale of tens of seconds. The dynamic localization observed is consistent with a spontaneous oscillation-type model which may be used in the essential positioning of replicated DNA prior to successful DNA segmentation into newly divided cells.
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