FoF1-ATP synthases in Escherichia coli (E. coli) bacteria are membrane-boundenzymes which use an internal proton-driven rotary double motor to catalyze thesynthesis of adenosine triphosphate (ATP). According to the 'chemiosmotichypothesis', a series of proton pumps generate the necessary pH difference plusan electric potential across the bacterial plasma membrane. These proton pumpsare redox-coupled membrane enzymes which are possibly organized insupercomplexes, as shown for the related enzymes in the mitochondrial innermembrane. We report diffusion measurements of single fluorescent FoF1-ATPsynthases in living E. coli by localization microscopy and single enzymetracking to distinguish a monomeric enzyme from a supercomplex-associated formin the bacterial membrane. For quantitative mean square displacement (MSD)analysis, the limited size of the observation area in the membrane with asignificant membrane curvature had to be considered. The E. coli cells had adiameter of about 500 nm and a length of about 2 to 3 \mum. Because the surfacecoordinate system yielded different localization precision, we applied asliding observation window approach to obtain the diffusion coefficient D =0.072 \mum2/s of FoF1-ATP synthase in living E. coli cells.
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