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

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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.


Contacts of IF3 with 30S Ribosomal SubunitThe CTD binds next to IF1 at the P site on top of h44 (position 1), and the NTD extends to the platform, while the linker lies on h23 and h24. Tyr75 (E. coli numbering) is also shown. See also Figure S4 and Movie S3.
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fig3: Contacts of IF3 with 30S Ribosomal SubunitThe CTD binds next to IF1 at the P site on top of h44 (position 1), and the NTD extends to the platform, while the linker lies on h23 and h24. Tyr75 (E. coli numbering) is also shown. See also Figure S4 and Movie S3.

Mentions: In tRNA-free structures (PICs 1A–1C), the dumbbell-shaped density is seen for nearly all of IF3 (Figure 3). The NTD binds near the 30S platform with a possible interaction with uS11; the bulk of the NTD is positioned away from 16S rRNA. The linker extends toward the P site lying on h23 (687–702) and h24 (787–792) (Escherichia coli numbering of 16S rRNA). The CTD density is observed at the top of h44 adjacent to the P site making interactions with both h44 (1494–1497), h24, and with G1517 of h45. The residues of IF3 involved in binding 16S rRNA (63–70, 86–97, 101–105) are mainly hydrophilic and basic. This position of IF3 on 30S agrees with the 16S rRNA cleavage in directed hydroxyl radical probing experiment of 30S⋅IF3 complex (Dallas and Noller, 2001). It is also consistent with protection of rRNA residues in h23 and h24 (G700, U701, G703, G791, and U793) by IF3 in chemical footprints (Muralikrishna and Wickstrom, 1989, Moazed et al., 1995) and other previous mutational studies (de Bellis et al., 1992, Tapprich et al., 1989). Tyr75 (E. coli numbering), a NTD/linker residue that when mutated reduces the fidelity of initiation (Maar et al., 2008), is in contact with the platform in these structures.


Large-Scale Movements of IF3 and tRNA during Bacterial Translation Initiation
Contacts of IF3 with 30S Ribosomal SubunitThe CTD binds next to IF1 at the P site on top of h44 (position 1), and the NTD extends to the platform, while the linker lies on h23 and h24. Tyr75 (E. coli numbering) is also shown. See also Figure S4 and Movie S3.
© Copyright Policy - CC BY
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5037330&req=5

fig3: Contacts of IF3 with 30S Ribosomal SubunitThe CTD binds next to IF1 at the P site on top of h44 (position 1), and the NTD extends to the platform, while the linker lies on h23 and h24. Tyr75 (E. coli numbering) is also shown. See also Figure S4 and Movie S3.
Mentions: In tRNA-free structures (PICs 1A–1C), the dumbbell-shaped density is seen for nearly all of IF3 (Figure 3). The NTD binds near the 30S platform with a possible interaction with uS11; the bulk of the NTD is positioned away from 16S rRNA. The linker extends toward the P site lying on h23 (687–702) and h24 (787–792) (Escherichia coli numbering of 16S rRNA). The CTD density is observed at the top of h44 adjacent to the P site making interactions with both h44 (1494–1497), h24, and with G1517 of h45. The residues of IF3 involved in binding 16S rRNA (63–70, 86–97, 101–105) are mainly hydrophilic and basic. This position of IF3 on 30S agrees with the 16S rRNA cleavage in directed hydroxyl radical probing experiment of 30S⋅IF3 complex (Dallas and Noller, 2001). It is also consistent with protection of rRNA residues in h23 and h24 (G700, U701, G703, G791, and U793) by IF3 in chemical footprints (Muralikrishna and Wickstrom, 1989, Moazed et al., 1995) and other previous mutational studies (de Bellis et al., 1992, Tapprich et al., 1989). Tyr75 (E. coli numbering), a NTD/linker residue that when mutated reduces the fidelity of initiation (Maar et al., 2008), is in contact with the platform in these structures.

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.