Clustering and dynamics of cytochrome bd‐I complexes in the Escherichia coli plasma membrane in vivo |
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| Authors: | Tchern Lenn Mark C Leake Conrad W Mullineaux |
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| Institution: | 1. School of Biological and Chemical Sciences, Queen Mary, University of London, Mile End Road, London E1 4NS, UK.;2. These authors contributed equally to the work.;3. Clarendon Laboratory, Department of Physics, University of Oxford, Parks Road, Oxford OX1 3PU, UK.;4. Oxford Centre for Integrative Systems Biology (OCISB), Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, UK. |
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| Abstract: | The cytochrome bd‐I complex of Escherichia coli is a respiratory terminal oxidase and an integral component of the cytoplasmic membrane. As with other respiratory components, the organization and dynamics of this complex in living membranes is unknown. We set out to visualize the distribution and dynamics of this complex in vivo. By exchanging cydB for cydB–gfpgcn4 on the E. coli chromosome, we produced a strain (YTL01) that expresses functional GFP‐tagged cytochrome bd‐I terminal oxidase complexes under wild‐type genetic control. We imaged live YTL01 cells using video‐rate epifluorescence and total internal reflection fluorescence (TIRF) microscopy in combination with fluorescence recovery after photobleaching (FRAP) and saw mobile spots of GFP fluorescence in plasma membranes. Numbers of GFP molecules per spot were quantified by step‐wise photobleaching giving a broad distribution with a mean of ～76, indicating that cytochrome bd‐I is concentrated in mobile patches in the E. coli plasma membrane. We hypothesize that respiration occurs in mobile membrane patches which we call ‘respirazones’. |
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