ABSTRACT
The essential ATP-binding cassette protein ABCE1 splits 80S ribosomes into 60S and 40S subunits after canonical termination or quality-control-based mRNA surveillance processes. However, the underlying splitting mechanism remains enigmatic. Here, we present a cryo-EM structure of the yeast 40S-ABCE1 post-splitting complex at 3.9-Å resolution. Compared to the pre-splitting state, we observe repositioning of ABCE1's iron-sulfur cluster domain, which rotates 150° into a binding pocket on the 40S subunit. This repositioning explains a newly observed anti-association activity of ABCE1. Notably, the movement implies a collision with A-site factors, thus explaining the splitting mechanism. Disruption of key interactions in the post-splitting complex impairs cellular homeostasis. Additionally, the structure of a native post-splitting complex reveals ABCE1 to be part of the 43S initiation complex, suggesting a coordination of termination, recycling, and initiation.
Subject(s)
ATP-Binding Cassette Transporters/chemistry , Ribosome Subunits, Small, Eukaryotic/chemistry , Saccharomyces cerevisiae Proteins/chemistry , Saccharomyces cerevisiae/enzymology , ATP-Binding Cassette Transporters/metabolism , ATP-Binding Cassette Transporters/ultrastructure , Cryoelectron Microscopy , Models, Biological , Models, Molecular , Peptide Chain Initiation, Translational , Peptide Chain Termination, Translational , Protein Binding , Ribosome Subunits, Small, Eukaryotic/metabolism , Ribosome Subunits, Small, Eukaryotic/ultrastructure , Saccharomyces cerevisiae/chemistry , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae Proteins/ultrastructureABSTRACT
Termination and ribosome recycling are essential processes in translation. In eukaryotes, a stop codon in the ribosomal A site is decoded by a ternary complex consisting of release factors eRF1 and guanosine triphosphate (GTP)-bound eRF3. After GTP hydrolysis, eRF3 dissociates, and ABCE1 can bind to eRF1-loaded ribosomes to stimulate peptide release and ribosomal subunit dissociation. Here, we present cryoelectron microscopic (cryo-EM) structures of a pretermination complex containing eRF1-eRF3 and a termination/prerecycling complex containing eRF1-ABCE1. eRF1 undergoes drastic conformational changes: its central domain harboring the catalytically important GGQ loop is either packed against eRF3 or swung toward the peptidyl transferase center when bound to ABCE1. Additionally, in complex with eRF3, the N-terminal domain of eRF1 positions the conserved NIKS motif proximal to the stop codon, supporting its suggested role in decoding, yet it appears to be delocalized in the presence of ABCE1. These results suggest that stop codon decoding and peptide release can be uncoupled during termination.
