Multi-Scale Simulations Provide Supporting Evidence for the Hypothesis of Intramolecular Protein Translocation in GroEL/GroES Complexes

The biological function of chaperone complexes is to assist the folding of non-native proteins. The widely studied GroEL chaperonin is a double-barreled complex that can trap non-native proteins in one of its two barrels. The ATP-driven binding of a GroES cap then results in a major structural change of the chamber where the substrate is trapped and initiates a refolding attempt. The two barrels operate anti-synchronously. The central region between the two barrels contains a high concentration of disordered protein chains, the role of which was thus far unclear. In this work we report a combination of atomistic and coarse-grained simulations that probe the structure and dynamics of the equatorial region of the GroEL/GroES chaperonin complex. Surprisingly, our simulations show that the equatorial region provides a translocation channel that will block the passage of folded proteins but allows the passage of secondary units with the diameter of an alpha-helix. We compute the free-energy barrier that has to be overcome during translocation and find that it can easily be crossed under the influence of thermal fluctuations. Hence, strongly non-native proteins can be squeezed like toothpaste from one barrel to the next where they will refold. Proteins that are already fairly close to the native state will not translocate but can refold in the chamber where they were trapped. Several experimental results are compatible with this scenario, and in the case of the experiments of Martin and Hartl, intra chaperonin translocation could explain why under physiological crowding conditions the chaperonin does not release the substrate protein.Author Summary: Chaperonin complexes capture proteins that have not yet reached their functional (“native”) state. Non-native proteins cannot perform their function correctly and threaten the survival of the cell. The chaperonins help these proteins to reach their native state. The prokaryotic GroEL-GroES chaperonin is an ellipsoidal protein complex that is approximately 16 nm long. It consists of two chambers that are joined at the bottom. Interestingly, protein repair by this chaperonin is not a one-step process. Typically, several capture and release steps are needed before the target protein reaches its native state. It is commonly assumed that substrate proteins cannot translocate, i.e., move inside the complex from one chamber to the other. In the absence of translocation, proteins that have not yet reached their functional conformation have to be released into the cytosol before being recaptured by a chaperonin. We present multi-scale simulations that show that it is, in fact, surprisingly easy for substrate proteins to translocate between the two chambers via an axial pore that is filled with disordered protein filaments. This finding suggests that non-native proteins can be squeezed like toothpaste from one chamber to the other: the incorrect structure of the protein is broken up during translocation and the protein has an increased probability to find its native state when it reaches the other chamber. The possibility for intra-chaperonin translocation obviates the need for a potentially dangerous release of non-native proteins.