Figure 6. In vivo initiation kinetics using Cy5 labeled initator tRNA^fMet.

a Simplified scheme displaying the predicted diffusional behavior of [Cy5]tRNA^fMet molecules being utilized on ribosomes. The dwell time of a functional [Cy5]tRNA^fMet binding event to a ribosome includes joining of the large ribosomal subunit and one cycle of peptide elongation. b MSD-estimated apparent diffusion coefficients of [Cy5]tRNA^fMet in DH5a cells (n = 31,498) treated with rifampicin (n = 8,389) or tetracycline (n = 21,259) injected. c HMM estimated occupancy of [Cy5]tRNA^fMet in the slow diffusional state (see Supplementary Table 1). The trajectories used were the same as in panel b. d MSD-estimated apparent diffusion coefficients of [Cy5]tRNA^fMet in DH5α cells overexpressing a short peptide with a rare Arg codon (AGG read by tRNA^Arg4, n = 19,900) or frequent Arg codon (CGU read by tRNA^Arg2, n = 25,236) in second position. e HMM estimated dwell times for [Cy5]tRNA^fMet in the slow state with or without short peptides overexpressed (see Supplementary Table 1). The trajectories used were the same as in panel d. Error bars in panel c and e represent bootstrap estimates of standard errors.