A Model for the Molecular Mechanism of an Engineered Light-Driven Protein Machine

Controllable protein-based machines and materials are of considerable interest for diverse biotechnological applications. We previously re-engineered an ATP-driven protein machine, a group II chaperonin, to function as a light-gated nanocage. Here we develop and test a model for the molecular mechanism of the re-engineered chaperonin, which undergoes a large-scale closed to open conformational change triggered by reversible photo-isomerization of a site-specifically attached azobenzene crosslinker. In silico experiments using all-atom simulations suggest that rigid body motions of protein subdomains couple the length changes of the crosslinker to rearrangements of the nucleotide-binding pocket, leading to cage opening. We tested this model by designing a mutant for which the orientation of the two protein subdomains forming the nucleotide-binding pocket is directly controlled by the crosslinker, and confirmed successful reversible photoswitching in vitro. The model probes the conformational cycle of group II chaperonins and offers a design principle for engineering other light-driven protein-based molecular machines.