Engineering the entropy-driven free-energy landscape of a dynamic nanoporous protein assembly

Abstract

De novo design and construction of stimuli-responsive protein assemblies that predictably switch between discrete conformational states remains an essential but highly challenging goal in biomolecular design. We previously reported synthetic, two-dimensional protein lattices self-assembled via disulfide bonding interactions, which endows them with a unique capacity to undergo coherent conformational changes without losing crystalline order. Here, we carried out all-atom molecular dynamics simulations to map the free-energy landscape of these lattices, validated this landscape through extensive structural characterization by electron microscopy and established that it is predominantly governed by solvent reorganization entropy. Subsequent redesign of the protein surface with conditionally repulsive electrostatic interactions enabled us to predictably perturb the free-energy landscape and obtain a new protein lattice whose conformational dynamics can be chemically and mechanically toggled between three different states with varying porosities and molecular densities.

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Fig. 1: Structural features of C98RhuA crystals.
Fig. 2: Thermodynamic analysis of C98RhuA lattice structural dynamics.
Fig. 3: Consequences of lattice compaction on solvent structure within the pore.
Fig. 4: Design and analysis of the CEERhuA construct.
Fig. 5: Chemical and mechanical switching behaviour of CEERhuA crystals.

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Acknowledgements

We thank M. Gilson, T. Kurtzman and S. Ramsey for helpful discussions regarding GIST, R. Subramanian for assistance with the generation of projection maps from computational models, and T. Baker for use of the electron microscopy facilities. This work was primarily supported by the US Department of Energy (Division of Materials Sciences, Office of Basic Energy Sciences; Award DE-SC0003844 to F.A.T.). F.P. was supported by the National Science Foundation through grant CHE-1453204 (computation). R.A. was supported in part by the University of California, San Diego NIH Molecular Biophysics Training Grant (T32-GM08326). All computer simulations were performed on the Extreme Science and Engineering Discovery Environment, which is supported by the National Science Foundation through grant ACI-1053575.

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R.A. co-initiated the project, designed and performed all of the experiments, simulations and data analysis, and co-wrote the paper. Y.S. performed the TEM data collection and analysis. F.P. guided the simulation design and computational data analysis, and co-wrote the paper. F.A.T. initiated the project, guided the experiment design and data analysis, and co-wrote the paper.

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Correspondence to Francesco Paesani or F. Akif Tezcan.

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Methods, Supplementary Figures 1–11, Supplementary Table 1 and Supplementary References

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Alberstein, R., Suzuki, Y., Paesani, F. et al. Engineering the entropy-driven free-energy landscape of a dynamic nanoporous protein assembly. Nature Chem 10, 732–739 (2018). https://doi.org/10.1038/s41557-018-0053-4

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