Single-shot characterization of enzymatic reaction constants Km and kcat by an acoustic-driven bubble-based fast micromixer

摘要

In this work we present an acoustofluidic approach for rapid, single-shot characterization of enzymatic reaction constants Km and kcat. The acoustofluidic design involves a bubble anchored in a horseshoe structure which can be stimulated by a piezoelectric transducer to generate vortices in the fluid. The enzyme and substrate can thus be mixed rapidly, within 100 ms, by the vortices to yield the product. Enzymatic reaction constants Km and kcat can then be obtained from the reaction rate curves for different concentrations of substrate while holding the enzyme concentration constant. We studied the enzymatic reaction for β-galactosidase and its substrate (resorufin β-D-galactopyranoside) and found Km and kcat to be 333±130 =M and 64±8 s⁻¹ respectively, which are in agreement with published data. Our approach is valuable for studying the kinetics of high-speed enzymatic reactions and other chemical reactions.