Toward a unifying mechanistic model of eukaryotic translation initiation through integrative single-molecule,...

During eukaryotic translation initiation, initiation factor proteins and the ribosomal small subunit undergo binding and dissociation reactions and conformational rearrangements that properly assemble a ribosome at the start codon of a messenger RNA. Building on extensive genetic and biochemical studies, single-molecule fluorescence experiments are revealing the time-dependent pathways of factor binding to, and dissociation from, the ribosomal small subunit and messenger RNA during initiation. Nonetheless, essential binding and/or dissociation events, conformational rearrangements, and the coupling between binding and conformational changes remain kinetically uncharacterized. Here, we summarize the status of single-molecule investigations of initiation and advocate for integrating single-molecule microscopy, structural biology, and molecular simulations to enable a time-dependent, molecular description of this fundamental step in gene expression.