Continuous Flow Reactors from Microfluidic Compartmentalization of Enzymes within Inorganic Microparticles

Abstract

Compartmentalization and selective transport of molecular species are key aspects of chemical transformations inside the cell. In an artificial setting, the immobilization of a wide range of enzymes onto surfaces is commonly used for controlling their functionality but such approaches can restrict their efficacy and expose them to degrading environmental conditions, thus reducing their activity. Here, we employ an approach based on droplet microfluidics to generate enzyme-containing microparticles that feature an inorganic silica shell that forms a semipermeable barrier. We show that this porous shell permits selective diffusion of the substrate and product while protecting the enzymes from degradation by proteinases and maintaining their functionality over multiple reaction cycles. We illustrate the power of this approach by synthesizing microparticles that can be employed to detect glucose levels through simultaneous encapsulation of two distinct enzymes that form a controlled reaction cascade. These results demonstrate a robust, accessible, and modular approach for the formation of microparticles containing active but protected enzymes for molecular sensing applications and potential novel diagnostic platforms.

Biomimetic Silica Encapsulation of Lipid Nanodiscs and β-Sheet-Stabilized Diacylglycerol Kinase

“…Two different silica-precipitating tags were fused to the BP-1 peptide: the full-length R5 peptide from the diatom C. fusiformis and a truncated version based on the C-terminal amino acids KRRIL only (R5′, Figure c). Approaches based on silaffin peptide directed biomimetic silica precipitation have been previously used to encapsulate biological structures such as eukaryotic cells and biomaterials as well as for the generation of protected enzymes and microcompartments , since their discovery more than 20 years ago. , Genetic manipulation to generate diatoms with new enzymatic functions based on their silica deposition machinery has also been reported as well as the use of synthetic silaffins to encapsulate soluble proteins. , Here, we aim for the encapsulation of a functional membrane protein in a fully controlled lipid or detergent environment without direct modification of the protein to allow transfer of this approach to other membrane proteins.…”

Let the Biocatalyst Flow

“…The microparticles were trapped in a microfluidic array device in which the enzyme activity could be tested in a single microparticle, which also provided information on reaction kinetic parameters and stability. 76…”

Toward the Development of Rapid, Specific, and Sensitive Microfluidic Sensors: A Comprehensive Device Blueprint

“…Owing to the increased surface to volume ratios of these bead-based reaction sites, the density of active receptors is much higher than those with planar surfaces. 137 It is worth noting that bead-based bioassay systems that incorporate packed bed or porous media rely on different transport mechanisms than those explained in this perspective. 138 However, the assumptions made in this perspective are still applicable in systems where the receptorcoated beads are immobilized in well-defined, closely packed patterns onto microchannel surfaces, with sufficient distance from the microchannel roofs.…”