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. Author manuscript; available in PMC: 2026 Jan 28.
Published in final edited form as: Nat Struct Mol Biol. 2025 Jul 28;32(11):2308–2318. doi: 10.1038/s41594-025-01629-y

Figure 2. Termination of transient eIF1 binding requires a translation start site and concentration-dependent binding by eIF5.

Figure 2.

(a) Cartoon schematic of the model mRNAs with either 50 nt (black), 200 nt (orange) long 5’UTRs or no AUG (purple). (b-e) Cumulative probability plots of the indicated eIF1 binding parameters observed on eIF5B-bound initiation complexes; the exceptions are the No AUG and TC-GDPNP data, which represent complexes that rarely or never progressed to eIF5B binding. Lines represent fits to exponential functions, and the derived mean times are reported on the plots. The 5’UTR length effects on eIF1 kinetics are contrasted in panel b. Panel c shows the effects of removing the AUG start site. Panel d contrasts use of TC-GTP against TC-GDPNP on β-globin mRNA. Panel e contrasts molar excess 290 nM eIF5 against a limiting concentration of 10 nM eIF5. For all conditions, 40 nM of Cy5-eIF1 and 290 nM eIF5 (unless as stated in panel e) were used. Panels b and c plot the same AUG (50 nt) data, and panels d and e plot the same β-globin data with 290 nM eIF5 and TC-GTP present. See Supplementary Table 1 for all rates, rate constants, and the number of complexes and binding events analyzed in each experiment.