A detailed molecular picture of protein folding during active translation

Abstract

All proteins can begin to fold on the ribosome, and many rely on co-translational folding to attain their native conformation. This process is not accounted for using structure prediction algorithms such as AlphaFold and its molecular details remain largely unknown. Here, we develop a hydrogen-deuterium pulse-labeling approach which reveals nascent polypeptide folding at a high level of structural detail and its kinetic coupling with translation. Two proteins exhibit hierarchical folding of structures smaller than a domain during translation. A third protein, however, does not have time to conformationally equilibrate on the ribosome, instead becoming kinetically trapped. This subsequently biases post-translational folding to avoid an aggregation-prone intermediate populated during refolding from denaturant. Our results reveal diverse strategies that promote robust protein folding during non-equilibrium translation.