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Updated: May 22, 2025
Toeprinting Analysis of Translation Initiation Complex Formation on Mammalian mRNAs
Published on: May 10, 2018
The translation inhibitors kasugamycin, edeine and GE81112 target distinct steps during 30S initiation complex
Haaris A Safdari1, Martino Morici1, Ana Sanchez-Castro2
1Institute for Biochemistry and Molecular Biology, University of Hamburg, 20146, Hamburg, Germany.
Abstract:
During bacterial translation initiation, the 30S ribosomal subunit, initiation factors, and initiator tRNA define the reading frame of the mRNA. This process is inhibited by kasugamycin, edeine and GE81112, however, their mechanisms of action have not been fully elucidated. Here we present cryo-electron microscopy structures of 30S initiation intermediate complexes formed in the presence of kasugamycin, edeine and GE81112 at resolutions of 2.0-2.9 Å. The structures reveal that all three antibiotics bind within the E-site of the 30S and preclude 30S initiation complex formation. While kasugamycin and edeine affect early steps of 30S pre-initiation complex formation, GE81112 stalls pre-initiation complex formation at a further step by allowing start codon recognition, but impeding IF3 departure. Collectively, our work highlights how chemically distinct compounds binding at a conserved site on the 30S can interfere with translation initiation in a unique manner.
Insights
Three antibiotics inhibit bacterial translation initiation by binding to the 30S ribosomal subunit's E-site. Kasugamycin, edeine, and GE81112 reveal distinct mechanisms impacting initiation complex formation and factor departure.
Area of Science:
- Molecular Biology
- Structural Biology
- Microbiology
Background:
- Bacterial translation initiation is crucial for protein synthesis, involving the 30S ribosomal subunit, initiation factors, and initiator tRNA.
- Kasugamycin, edeine, and GE81112 are known inhibitors of this process, but their precise mechanisms remain unclear.
Purpose of the Study:
- To elucidate the molecular mechanisms by which kasugamycin, edeine, and GE81112 inhibit bacterial translation initiation.
- To determine the structural basis of antibiotic interaction with the 30S ribosomal initiation complex.
Main Methods:
- Cryo-electron microscopy (cryo-EM) was used to determine high-resolution structures (2.0-2.9 Å) of 30S initiation intermediate complexes.
- These complexes were formed in the presence of kasugamycin, edeine, and GE81112.
Main Results:
- All three antibiotics were found to bind within the E-site of the 30S subunit, preventing 30S initiation complex formation.
- Kasugamycin and edeine interfere with early stages of 30S pre-initiation complex assembly.
- GE81112 allows start codon recognition but blocks the departure of initiation factor IF3, stalling the complex formation at a later step.
Conclusions:
- Chemically distinct antibiotics can inhibit bacterial translation initiation through unique mechanisms by binding to a conserved site on the 30S ribosomal subunit.
- Understanding these mechanisms provides insights into novel antibacterial strategies targeting protein synthesis.
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