Cells trigger the reversible formation of multi-component protein assemblies in response to their metabolic state and external stimuli. These dynamic assemblies confer several functional benefits, including enhancement and optimization of metabolic pathways. Assembly formation occurs through triggerable, highly specific electrostatic and hydrophobic interactions between proteins, resulting in structure formation on the micrometer scale. Development of synthetic, stimulus-responsive protein assemblies, whose formation is controlled by non-invasive external stimuli such as light, would allow for the interrogation of the underlying physicochemical design principles of protein-based assemblies, and enable several applications in synthetic biology. We have developed a multi-scale modular method for computationally designing and modeling photoswitchable proteins that self-assemble in response to light. The design process involves atomic-resolution modeling of fusions of oligomeric proteins with hetero-dimeric coiled coil peptides that have been appropriately labeled with photoresponsive azobenzene dyes. Multivalent and reversible photocontrolled protein-protein interface formation is expected to cause assembly formation on the micrometer scale. Here we demonstrate that designed proteins can bind and assemble, and that point mutation-induced alterations to their individual structures on the sub-nanometer scale predictively determine supramolecular structure formation on the micrometer scale. To probe the mechanistic aspects of the assembly process, we use Brownian Dynamics simulations of coarse grained protein models. These simulations provide insight into the specific interactions governing assembly formation and aggregation dynamics. The self-assembled structures observed in simulations are in good agreement with experimental observations using microscopy techniques. We also demonstrate successful labeling of these proteins with azobenzene, a chemical photoswitch, making the designed protein assembly light responsive. The described design approach is general, and should enable in the bottom-up design and modeling of diverse photocontrolled protein assemblies with a variety of protein components.

An integrated computational-experimental approach for designing photoswitchable protein assemblies

256th National Meeting (2018)

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