title: Single-shot characterization of enzymatic reaction constants Km and kcat by an acoustic-driven, bubble-based fast micromixer.
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.
authors.affiliation: Department of Chemical Engineering, The Pennsylvania State University, University Park, Pennsylvania 16802, United States.
authors.forename: Yuliang
authors.initials: Y
authors.lastname: Xie
authors.forename: Daniel
authors.initials: D
authors.lastname: Ahmed
authors.forename: Michael Ian
authors.initials: MI
authors.lastname: Lapsley
authors.forename: Sz-Chin Steven
authors.initials: SC
authors.lastname: Lin
authors.forename: Ahmad Ahsan
authors.initials: AA
authors.lastname: Nawaz
authors.forename: Lin
authors.initials: L
authors.lastname: Wang
authors.forename: Tony Jun
authors.initials: TJ
authors.lastname: Huang
mesh_terms: D000162:Acoustics; D004129:Dimethylpolysiloxanes / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology; D005697:Galactosides / Q000378:metabolism; D007700:Kinetics; D046210:Microfluidic Analytical Techniques; D010078:Oxazines / Q000378:metabolism; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D004129:Dimethylpolysiloxanes; D005697:Galactosides; D010078:Oxazines; C013830:baysilon; C056561:resorufin galactopyranoside; D001616:beta-Galactosidase