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. 2011 Nov 17;40(5):2054–2064. doi: 10.1093/nar/gkr1031

Figure 4.

Figure 4.

Fast kinetics measurements of the steps of initiation. The time course of association of MRE600 (black trace), MG1655 (blue trace) (both with two L12 dimers) and JE105 (red trace) (with single L12 dimer) 50S subunits with naked 30S subunits (a), or with 30S preIC containing mRNA, fMet-tRNAfMet, IF1 and IF2·GTP (b) followed in stopped-flow by monitoring increase in light scattering at 430 nm. The insert in (b) shows the same reaction for prolonged time. (c) The plots showing linear dependence of the observed rates of subunit association on 50S concentration in reactions with 30S preIC. (d) Single round Pi release with MRE600 (black trace) and JE105 (red trace) ribosomes, measured in parallel to subunit association in the same reaction as in (b). The increase in MDCC fluorescence upon Pi binding to PBP-MDCC was monitored at 464 nm (λEx = 425 nm) (e) Schematic representation of translation initiation where average times for individual steps were estimated for two L12 dimer MRE600 (in black) and single L12 dimer JE105 (in red) ribosomes (see ‘Materials and Methods’ section for details).