Tuning in-meso-crystallized lysozyme polymorphism by lyotropic liquid crystal symmetry

Lipid-based lyotropic liquid crystals (LLCs) show great potential for applications in fields as diverse as food technology, cosmetics, pharmaceutics, or structural biology. Recently, these systems have provided a viable alternative to the difficult process of membrane protein crystallization, owing to their similarities with cell membranes. Nonetheless, the process of in-meso crystallization of proteins still remains poorly understood. In this study, we demonstrate that in-meso crystal morphologies of lysozyme (LSZ), a model hydrophilic protein, can be controlled by both the composition and symmetry of the mesophase, inferring a possible general influence of the LLC space group on the protein crystal polymorphism. Lysozyme was crystallized in-meso from three common LLC phases (lamellar, inverse hexagonal, and inverse bicontinuous cubic) composed of monolinolein and water. Different mixing ratios of mesophase to crystallization buffer were used in order to tune crystallization both in the bulk mesophase and in excess water conditions. Two distinct mechanisms of crystallization were shown to take place depending on available water in the mesophases. In the bulk mesophases, protein nuclei form and grow within structural defects of the mesophase and partially dehydrate the system inducing order-to-order transitions of the liquid crystalline phase toward stable symmetries in conditions of lower hydration. The formed protein crystals eventually macrophase separate from the mesophase allowing the system to reach its final symmetry. On the other hand, when excess water is available, protein molecules diffuse from the water channels into the excess water, where the crystallization process can take place freely, and with little to no effect on the structure and symmetry of the lyotropic liquid crystals