Biomimetic Active Protein Droplets
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2022
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Doctoral Thesis
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yes
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Abstract
Living cells are continuously driven out of equilibrium by the intricate interplay of the many chemical processes that we call life. They are able to sustain these processes by harvesting energy from their environment through metabolism, which extracts potential energy from a chemical fuel. These reactions happen in a cellular environment that is a crowded, complex and highly dynamic mixture of concentrated macromolecules, oligomers and small molecules. For this reasons, describing and understanding biological phenomena is extremely challenging, and in vitro experiments are typically performed in very dilute, close-to-equilibrium conditions, which fail to capture the essential features ofthe cellular medium. In this thesis, we introduce a minimal system that operates at similar metabolic densities, protein concentrations and length scales as living cells, while still being simple enough to understand and control. Inspired by membraneless organelles, we exploit the liquid-liquid phase separation of bovine serum albumin in presence of a polymer to create highly crowded protein domains, which spontaneously partition enzymes. By dispersing the droplets in a reservoir ofsubstrate-loaded buffer, we are able to obtain local metabolic densities that match those of the hungriest living organisms. The membraneless nature ofthese microreactors allows efficient exchange of chemicals and heat, and in turn the possibility to achieve steady states. We further demonstrate that metabolic activity is capable of promoting a plethora ofbiomimetic behaviors, like the formation of pH gradients, microscopic flows, and droplet motion. Our work provides a flexible platform for the study of biological phenomena in an environment that mimics the cellular cytoplasm, and more in general for the understanding of systems driven far from equilibrium and their emergent collective behavior. It also establishes a framework and a set of design rules for working with active droplet systems, including the development of a novel slippery glass coating that exhibits zero-wetting conditions for a wide range ofmacromolecular condensates and lipid vesicles.
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ETH Zurich
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09573 - Dufresne, Eric (ehemalig) / Dufresne, Eric (former)