Functional mapping of the E. coli E. coli translational machinery using single‐molecule tracking

摘要

Summary The organization of the chromosomal DNA and ribosomes in living Escherichia coli is compared under two growth conditions: ‘fast’ (50?min doubling time) and ‘slow’ (147?min doubling time). Superresolution fluorescence microscopy reveals strong DNA‐ribosome segregation in both cases. In both fast and slow growth, free ribosomal subunits evidently must circulate between the nucleoid (where they initiate co‐transcriptional translation) and ribosome‐rich regions (where most translation occurs). Single‐molecule diffusive behavior dissects the ribosome copies into translating 70S polysomes and free 30S subunits, providing separate spatial distributions for each. In slow growth, ~21,000 total 30S copies/cell comprise ~65% translating 70S ribosomes and ~35% free 30S subunits. The ratio of 70S ribosomes to free 30S subunits is ~2.5 outside the nucleoid and ~0.50 inside the nucleoid. This new level of quantitative detail may motivate development of comprehensive, three‐dimensional reaction‐diffusion models of ribosome, DNA, mRNA and RNAP spatial distributions and dynamics within the E. coli cytoplasm.