(165l) Understanding Membrane Permeability of Proteinosomes Self-Assembled from Globular Fusion Proteins

To better understand the complexity of life on a microscopic level, biological processes can be simplified in engineered materials based on bottom-up construction of biomimetic protocells. Although liposomes and polymersomes have been developed for decades as synthetic cell platforms, protein-assembled vesicles, called proteinosomes, have recently attracted attention to utilize functional proteins in the grand challenge of synthetic cell platforms. We engineer globular protein vesicles (GPVs), which are self-assembled from recombinant fusion proteins including full-sized, globular domains, for synthetic cell development. GPV’s offer significant advantages not yet realized in existing protocell platforms, namely the highly variable integration of functional proteins and precise control over protein orientation. Although GPVs offer promising benefits to existing protocell platforms, their novelty in the field necessitates further understanding of their intrinsic properties, including selective permeability, stimuli-responsiveness, and fusion/fission. Herein, we focus on understanding of membrane permeability in GPVs to regulate metabolic cascade reactions. We hypothesized that membrane permeability can be directly manipulated through thermal gating of GPV membranes owing to lowest critical solution temperature (LCST) behavior of the elastin like polypeptide (ELP) building blocks. We expect temperature dependent conformational changes of ELP hydrophobic tails to cause relaxation or tightening of the vesicle membrane, altering the permeability via temperature control. Selective permeability will be investigated through diffusion of different sized molecules with fluorescent tags at various temperatures. Demonstration of this mechanism would allow for active control of diffusion dependent reactions found in biological systems like metabolic cascade reactions, and signal amplification.