Human Ebp1 is a member of the proliferation-associated 2G4 (PA2G4) family and plays an important role in cancer regulation. Ebp1 shares the methionine aminopeptidase (MetAP) fold and binds to mature 80S ribosomes for translational control. Here, we present a cryo-EM single particle analysis reconstruction of Ebp1 bound to non-translating human 80S ribosomes at a resolution range from 3.3 to~8 Å. Ebp1 blocks the tunnel exit with major interactions to the general uL23/uL29 docking site for nascent chain-associated factors complemented by eukaryote-specific eL19 and rRNA helix H59. H59 is defined as dynamic adaptor undergoing significant remodeling upon Ebp1 binding. Ebp1 recruits rRNA expansion segment ES27L to the tunnel exit via specific interactions with rRNA consensus sequences. The Ebp1-ribosome complex serves as a template for MetAP binding and provides insights into the structural principles for spatial coordination of co-translational events and molecular triage at the ribosomal tunnel exit.

SummaryRibosomes are among the most abundant and complex machineries in the cell, however, the turnover of their subunits remains poorly understood. Here, we apply proteomic flux and cryo-electron microscopy analyses to interrogate the ribosome life cycle in human cells. We show that subpopulations of ribosomal subunits coexist, which vary in turnover kinetics and structure. Specifically, 80S ribosomes have a much longer half-life than free 40S and 60S ribosomal subunits, indicating that they represent distinct subunit pools that poorly intermix. Translation inhibition starkly increases the pool-size of 80S ribosomes in a translationally idle state and induces ribophagy of old ribosomes, ultimately rejuvenating the ribosome fleet. Our findings provide a comprehensive model for ribosome turnover and its regulation via translational activity.

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