Abstract
Bacterial translation initiation factor IF2 promotes ribosomal subunit association, recruitment, and binding of fMet-tRNA to the ribosomal P-site and initiation dipeptide formation. Here, we present the solution structures of GDP-bound and apo-IF2-G2 of Bacillus stearothermophilus and provide evidence that this isolated domain binds the 50 S ribosomal subunit and hydrolyzes GTP. Differences between the free and GDP-bound structures of IF2-G2 suggest that domain reorganization within the G2-G3-C1 regions underlies the different structural requirements of IF2 during the initiation process. However, these structural signals are unlikely forwarded from IF2-G2 to the C-terminal fMet-tRNA binding domain (IF2-C2) because the connected IF2-C1 and IF2-C2 modules show completely independent mobility, indicating that the bacterial interdomain connector lacks the rigidity that was found in the archaeal IF2 homolog aIF5B.
Highlights
A central question in translation initiation is how GTPase activity and fMet-tRNA positioning are connected
Our results demonstrate that the B. stearothermophilus IF2-G21⁄7GTP complex can weakly bind the B. stearothermophilus 50 S ribosomal subunits but not the 30 S subunits
We conclude that IF2-G2 houses the functional elements responsible for GTP binding, the catalytic center for GTP hydrolysis, and at least part of the region for establishing functional interactions with the 50 S ribosomal subunit, it does not contain the region responsible for the interaction with the 30 S subunit
Summary
A central question in translation initiation is how GTPase activity and fMet-tRNA positioning are connected. Differences between the free and GDP-bound structures of IF2-G2 suggest that domain reorganization within the G2-G3-C1 regions underlies the different structural requirements of IF2 during the initiation process These structural signals are unlikely forwarded from IF2-G2 to the C-terminal fMet-tRNA binding domain (IF2-C2) because the connected IF2-C1 and IF2-C2 modules show completely independent mobility, indicating that the bacterial interdomain connector lacks the rigidity that was found in the archaeal IF2 homolog aIF5B. An elongated shape in bacterial IF2 would allow the simultaneous placement of the fMettRNA anticodon stem loop in the ribosomal 30 S P-site and the acceptor end in the 50 S peptidyltransferase center Despite their structural similarity, substantial deviations exist between bacterial IF2 and archaeal/eukaryal a/eIF5B functions. The results demonstrate that the behavior of bacterial IF2 is substantially different from its archaeal homolog aIF5B
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