Fabrication of Hierarchical Bioinspired Superstructures Bearing Different Charges and Tunable Ability to Promote Protein Crystallization

Fabrication of functional anisotropic polymeric nanostructures is of great interest and enables a wide range of applications such as protein crystallization. In this sense, we prepared a series of supramolecular nanostructures bearing different surface charges, including star-like superstructures and two-dimensional (2D) nanosheets from diblock copolymers containing crystalline polypeptoids via a hierarchical assembly strategy in a one-pot preparation. We demonstrate that the star-like structures possess a central core with less ordered polypeptoid chains and crystalline emanated branches with oriented molecular packing by grazing incidence wide-angle X-ray scattering (GIWAXS). Further, we developed a new strategy to apply the obtained polypeptoid-based superstructures for the promotion of the nucleation and growth of four types of protein crystals over a wide range of isoelectric points. To the best of our knowledge, this is the first report utilizing protein-mimetic supramolecular structures to efficiently promote the growth of protein crystals. Notably, the protein crystal quantity and quality are greatly improved by the addition of supramolecular nanostructures. In addition, the heterogeneous nucleation has been greatly enhanced, as indicated by the increased heterogeneous nucleation rate (Nhetero). The maximum Nhetero can reach up to 14 with the amine-containing nanostructures, larger than previously reported values. We demonstrate that protein crystal growth is highly dependent on the surface charge of the nanostructures. Such a broadly applicable approach presents great potential in protein crystallization for biotechnological applications.