Chain Length-Dependent Conformations of Co-Translational Folding Intermediates of P22 Tailspike

Newly synthesized proteins must form their native structure in the crowded environment of the cell, while avoiding non-native conformations that can lead to aggregation. P22 tailspike is a homotrimer prone to aggregation via misfolding of its central beta-helix domain in vitro. To assess whether co-translational folding enables newly synthesized tailspike chains to avoid aggregation-prone conformations in vivo, a novel method was first developed to produce stalled ribosome nascent chain complexes. This new method was used to measure anti-tailspike monoclonal antibody binding to and partial protease digestion of four different tailspike nascent chain lengths. These experiments reveal ribosome-bound nascent tailspike chains populate ordered conformations with some native-state structural features, but these conformations are distinct from the predominant conformations of tailspike in vitro refolding intermediates and refolded, unstalled tailspike truncations. These results suggest the aggregation-prone beta-helix domain pre-organizes co-translationally, prior to chain release, and that this conformation is distinct from the global energy minimum for the truncated free chain in solution.