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Abstract Directed evolution generates novel biomolecules with desired functions by iteratively diversifying the genetic sequence of wildtype biomolecules, relaying the genetic information to the molecule with function, and selecting the variants that progresses towards the properties of interest. While traditional directed evolution consumes significant labor and time for each step, continuous evolution seeks to automate all steps so directed evolution can proceed with minimum human intervention and dramatically shortened time. A major application of continuous evolution is the generation of novel enzymes, which catalyze reactions under conditions that are not favorable to their wildtype counterparts, or on altered substrates. The challenge to continuously evolve enzymes lies in automating sufficient, unbiased gene diversification, providing selection for a wide array of reaction types, and linking the genetic information to the phenotypic function. Over years of development, continuous evolution has accumulated versatile strategies to address these challenges, enabling its use as a general tool for enzyme engineering. As the capability of continuous evolution continues to expand, its impact will increase across various industries. In this review, we summarize the working mechanisms of recently developed continuous evolution strategies, discuss examples of their applications focusing on enzyme evolution, and point out their limitations and future directions.more » « less
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Abstract Dynamic microcapsules are reported that exhibit shell membranes with fast and reversible changes in permeability in response to external stimuli. A hydrophobic anhydride monomer is employed in the thiol–ene polymerization as a disguised precursor for the acid‐containing shells; this enables the direct encapsulation of aqueous cargo in the liquid core using microfluidic fabrication of water‐in‐oil‐in‐water double emulsion drops. The poly(anhydride) shells hydrolyze in their aqueous environment without further chemical treatment, yielding cross‐linked poly(acid) microcapsules that exhibit trigger‐responsive and reversible property changes. The microcapsule shell can actively be switched numerous times between impermeable and permeable due to the exceptional mechanical properties of the thiol–ene network that prevent rupture or failure of the membrane, allowing it to withstand the mechanical stresses imposed on the capsule during the dynamic property changes. The permeability and molecular weight cutoff of the microcapsules can dynamically be controlled with triggers such as pH and ionic environment. The reversibly triggered changes in permeability of the shell exhibit a response time of seconds, enabling actively adjustable release profiles, as well as on‐demand capture, trapping, and release of cargo molecules with molecular selectivity and fast on‐off rates.more » « less
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Abstract Nanoparticle‐shelled bubbles, prepared with glass capillary microfluidics, are functionalized to produce catalytic micromotors that exhibit novel assembly and disassembly behaviors. Stable microbubble rafts are assembled at an air–solvent interface of nonaqueous propylene carbonate (PC) solvent by creating a meniscus using a glass capillary. Upon the addition of hydrogen peroxide fuel, catalytic microbubbles quickly break free from the bubble raft by repelling from each other and self‐propelling at the air–fuel interface (a mixture of PC and aqueous hydrogen peroxide). While most of micromotors generate oxygen bubbles on the outer catalytic shell, some micromotors contain cracks and eject bubbles from the hollow shells containing air. Nanoparticle‐shelled bubbles with a high buoyancy force are particularly attractive for studying novel propulsion modes and interactions between catalytic bubble micromotors at air–fuel interfaces.more » « less
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Abstract The suppression of lithium dendrite is critical to the realization of lithium metal batteries. 3D conductive framework, among different approaches, has shown very promising results in dendrite suppression. A novel cost‐effective and versatile dip‐coating method is presented here to make 3D conductive framework. Various substrates with different geometries are coated successfully with copper, including electrically insulating glass fiber (GF) or rice paper and conducting Ni foam. In particular, the as‐prepared copper coated GF shows promising results to serve as the lithium metal substrate by the electrochemical battery tests. The method significantly broadens the candidate materials database for 3D conductive framework to include all kinds of intrinsically insulating 3D substrates.more » « less
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