Magnetically Directed Two-Dimensional Crystallization of OmpF Membrane Proteins in Block Copolymers

Two-dimensional (2D) alignment and crystallization of membrane proteins (MPs) is increasingly important in characterizing their three-dimensional (3D) structure, in designing pharmacological agents, and in leveraging MPs for biomimetic devices. Large, highly ordered MP 2D crystals in block copolymer (BCP) matrices are challenging to fabricate, but a facile and scalable technique for aligning and crystallizing MPs in thin-film geometries would rapidly translate into applications. This work introduces a novel method to grow larger and potentially better ordered 2D crystals by performing the crystallization process in the presence of a strong magnetic field. We demonstrate the efficacy of this approach using a beta-barrel MP, outer membrane protein F (OmpF), in short-chain polybutadiene-poly(ethylene oxide) (PB-PEO) membranes. Crystals grown in a magnetic field were up to 5 times larger than conventionally grown crystals, and a signal-to-noise (SNR) analysis of diffraction peaks in Fourier transforms of specimens imaged by negative-stain electron microscopy (EM) and cryo-EM showed twice as many high-SNR diffraction peaks, indicating that the magnetic field also improves crystal order.