Article

Single-Shot Characterization of Enzymatic Reaction Constants K-m and k(cat) by an Acoustic-Driven, Bubble-Based Fast Micromixer

Department of Chemical Engineering, The Pennsylvania State University , University Park, Pennsylvania 16802, United States.
Analytical Chemistry (Impact Factor: 5.83). 07/2012; 84(17):7495-501. DOI: 10.1021/ac301590y
Source: PubMed

ABSTRACT In this work we present an acoustofluidic approach for rapid, single-shot characterization of enzymatic reaction constants K(m) and k(cat). 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 K(m) and k(cat) 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 K(m) and k(cat) to be 333 ± 130 μM 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.

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