Cryo-EM of Ribosomal 80S Complexes with Termination Factors Reveals the Translocated Cricket Paralysis Virus IRES
2015
Muhs, Margarita | Hilal, Tarek | Mielke, Thorsten | Skabkin, Maxim A. | Sanbonmatsu, Karissa Y. | Pestova, Tatyana V. | Spahn, Christian M.T.
The cricket paralysis virus (CrPV) uses an internal ribosomal entry site (IRES) to hijack the ribosome. In a remarkable RNA-based mechanism involving neither initiation factor nor initiator tRNA, the CrPV IRES jumpstarts translation in the elongation phase from the ribosomal A site. Here, we present cryoelectron microscopy (cryo-EM) maps of 80S⋅CrPV-STOP⋅eRF1⋅eRF3⋅GMPPNP and 80S⋅CrPV-STOP⋅eRF1 complexes, revealing a previously unseen binding state of the IRES and directly rationalizing that an eEF2-dependent translocation of the IRES is required to allow the first A-site occupation. During this unusual translocation event, the IRES undergoes a pronounced conformational change to a more stretched conformation. At the same time, our structural analysis provides information about the binding modes of eRF1⋅eRF3⋅GMPPNP and eRF1 in a minimal system. It shows that neither eRF3 nor ABCE1 are required for the active conformation of eRF1 at the intersection between eukaryotic termination and recycling.
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