The ribosome synchronizes folding and assembly to promote oligomeric protein biogenesis

Natural proteins often form intricate multidomain, oligomeric architectures. This presents a prima facie challenge to cellular homeostasis, as topologically complex proteins seldom refold efficiently in vitro. Here, we show that the efficient folding and assembly of the five-domain homotetramer β-galactosidase is obligatorily coupled to its synthesis on the ribosome, and we define the underlying mechanisms. During refolding from a denaturant, maturation of the catalytic domain is frustrated. Assembly outpaces monomer folding, and non-native oligomers accumulate. Efficient de novo folding is characterized by segmental domain folding, shaped by the binding of a nascent amphipathic helix to a cryptic pocket on uL23 on the ribosome surface. Homomer assembly also initiates cotranslationally via recruitment of a full-length subunit to the nascent polypeptide, and the failure to do so results in misassembly. Our results reveal how the ribosome can dictate the timing of folding and assembly to enable efficient biogenesis of a topologically complex protein.