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Large-Scale Movements of IF3 and tRNA during Bacterial Translation Initiation

View Article: PubMed Central - PubMed

ABSTRACT

In bacterial translational initiation, three initiation factors (IFs 1–3) enable the selection of initiator tRNA and the start codon in the P site of the 30S ribosomal subunit. Here, we report 11 single-particle cryo-electron microscopy (cryoEM) reconstructions of the complex of bacterial 30S subunit with initiator tRNA, mRNA, and IFs 1–3, representing different steps along the initiation pathway. IF1 provides key anchoring points for IF2 and IF3, thereby enhancing their activities. IF2 positions a domain in an extended conformation appropriate for capturing the formylmethionyl moiety charged on tRNA. IF3 and tRNA undergo large conformational changes to facilitate the accommodation of the formylmethionyl-tRNA (fMet-tRNAfMet) into the P site for start codon recognition.

No MeSH data available.


IF3 on 30S, Related to Figures 3 and 4(A) The CTD in Position1 would clash with fMet-tRNAfMet in a previously reported 30S initiation complex (Simonetti et al., 2008). Only one fMet-tRNAfMet is shown for clarity, as a similar clash is also observed in a subsequent study (Julián et al., 2011).(B) The CTD in Position1 (orange) and Position2 (red) has strong clashes with H69 of large ribosomal subunit (blue) in 70S. The NTD however has no steric hindrance with 50S.(C) The CTD in Position1 (cyan ribbon) and Position1′ (orange surface) would clash with fMet-tRNAfMet (green surface) in PIC-4.(D) Superposition of IF3 in PIC-1C (yellow), PIC-2A (cyan) and PIC-3 (orange) highlighting the movements of the linker (between Asp61) and NTD. The tRNA in PIC-3 is shown. The linker in PIC-3 makes an angle of about 50 degrees with the original position. The NTD moves 28 Å compared to PIC-2A and about 38 Å compared to PIC-1C.(E) Superposition of the CTD of IF3 in PIC-1C (yellow), PIC-2A (cyan), PIC-2B (green), PIC-2C (purple), PIC-3 (orange), PIC-4 (red) highlights the movements of its NTD with respect to it.
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figs4: IF3 on 30S, Related to Figures 3 and 4(A) The CTD in Position1 would clash with fMet-tRNAfMet in a previously reported 30S initiation complex (Simonetti et al., 2008). Only one fMet-tRNAfMet is shown for clarity, as a similar clash is also observed in a subsequent study (Julián et al., 2011).(B) The CTD in Position1 (orange) and Position2 (red) has strong clashes with H69 of large ribosomal subunit (blue) in 70S. The NTD however has no steric hindrance with 50S.(C) The CTD in Position1 (cyan ribbon) and Position1′ (orange surface) would clash with fMet-tRNAfMet (green surface) in PIC-4.(D) Superposition of IF3 in PIC-1C (yellow), PIC-2A (cyan) and PIC-3 (orange) highlighting the movements of the linker (between Asp61) and NTD. The tRNA in PIC-3 is shown. The linker in PIC-3 makes an angle of about 50 degrees with the original position. The NTD moves 28 Å compared to PIC-2A and about 38 Å compared to PIC-1C.(E) Superposition of the CTD of IF3 in PIC-1C (yellow), PIC-2A (cyan), PIC-2B (green), PIC-2C (purple), PIC-3 (orange), PIC-4 (red) highlights the movements of its NTD with respect to it.

Mentions: The CTD in these complexes is positioned such that it would clash with the tRNA in reported structures of the 30S PIC (Simonetti et al., 2008, Julián et al., 2011; Figure S4A). This P-site location of the CTD (henceforth termed as position 1) features several highly conserved interactions with IF1 as well as with mRNA in the P site. CTD at position 1 also occludes the binding site for H69 of large ribosomal subunit and hence would prevent premature subunit association (Figure S4B).


Large-Scale Movements of IF3 and tRNA during Bacterial Translation Initiation
IF3 on 30S, Related to Figures 3 and 4(A) The CTD in Position1 would clash with fMet-tRNAfMet in a previously reported 30S initiation complex (Simonetti et al., 2008). Only one fMet-tRNAfMet is shown for clarity, as a similar clash is also observed in a subsequent study (Julián et al., 2011).(B) The CTD in Position1 (orange) and Position2 (red) has strong clashes with H69 of large ribosomal subunit (blue) in 70S. The NTD however has no steric hindrance with 50S.(C) The CTD in Position1 (cyan ribbon) and Position1′ (orange surface) would clash with fMet-tRNAfMet (green surface) in PIC-4.(D) Superposition of IF3 in PIC-1C (yellow), PIC-2A (cyan) and PIC-3 (orange) highlighting the movements of the linker (between Asp61) and NTD. The tRNA in PIC-3 is shown. The linker in PIC-3 makes an angle of about 50 degrees with the original position. The NTD moves 28 Å compared to PIC-2A and about 38 Å compared to PIC-1C.(E) Superposition of the CTD of IF3 in PIC-1C (yellow), PIC-2A (cyan), PIC-2B (green), PIC-2C (purple), PIC-3 (orange), PIC-4 (red) highlights the movements of its NTD with respect to it.
© Copyright Policy - CC BY
Related In: Results  -  Collection

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figs4: IF3 on 30S, Related to Figures 3 and 4(A) The CTD in Position1 would clash with fMet-tRNAfMet in a previously reported 30S initiation complex (Simonetti et al., 2008). Only one fMet-tRNAfMet is shown for clarity, as a similar clash is also observed in a subsequent study (Julián et al., 2011).(B) The CTD in Position1 (orange) and Position2 (red) has strong clashes with H69 of large ribosomal subunit (blue) in 70S. The NTD however has no steric hindrance with 50S.(C) The CTD in Position1 (cyan ribbon) and Position1′ (orange surface) would clash with fMet-tRNAfMet (green surface) in PIC-4.(D) Superposition of IF3 in PIC-1C (yellow), PIC-2A (cyan) and PIC-3 (orange) highlighting the movements of the linker (between Asp61) and NTD. The tRNA in PIC-3 is shown. The linker in PIC-3 makes an angle of about 50 degrees with the original position. The NTD moves 28 Å compared to PIC-2A and about 38 Å compared to PIC-1C.(E) Superposition of the CTD of IF3 in PIC-1C (yellow), PIC-2A (cyan), PIC-2B (green), PIC-2C (purple), PIC-3 (orange), PIC-4 (red) highlights the movements of its NTD with respect to it.
Mentions: The CTD in these complexes is positioned such that it would clash with the tRNA in reported structures of the 30S PIC (Simonetti et al., 2008, Julián et al., 2011; Figure S4A). This P-site location of the CTD (henceforth termed as position 1) features several highly conserved interactions with IF1 as well as with mRNA in the P site. CTD at position 1 also occludes the binding site for H69 of large ribosomal subunit and hence would prevent premature subunit association (Figure S4B).

View Article: PubMed Central - PubMed

ABSTRACT

In bacterial translational initiation, three initiation factors (IFs 1–3) enable the selection of initiator tRNA and the start codon in the P site of the 30S ribosomal subunit. Here, we report 11 single-particle cryo-electron microscopy (cryoEM) reconstructions of the complex of bacterial 30S subunit with initiator tRNA, mRNA, and IFs 1–3, representing different steps along the initiation pathway. IF1 provides key anchoring points for IF2 and IF3, thereby enhancing their activities. IF2 positions a domain in an extended conformation appropriate for capturing the formylmethionyl moiety charged on tRNA. IF3 and tRNA undergo large conformational changes to facilitate the accommodation of the formylmethionyl-tRNA (fMet-tRNAfMet) into the P site for start codon recognition.

No MeSH data available.