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Updated: Jun 24, 2026

Hot Biological Catalysis: Isothermal Titration Calorimetry to Characterize Enzymatic Reactions
Published on: April 4, 2014
Determination of kinetic parameters, Km and kcat, with a single experiment on a chip
Sachin Jambovane1, Evert C Duin, Se-Kwon Kim
1Materials Research and Education Center, Department of Mechanical Engineering, Auburn University, Auburn, Alabama 36849, USA.
Abstract:
We have demonstrated a multistep enzyme reaction on a chip to determine the key kinetic parameters of enzyme reaction. We designed and fabricated a fully integrated microfluidic chip to have sample metering, mixing, and incubation functionalities. The chip generates a gradient of reagent concentrations in 11 parallel processors. We used beta-galactosidase and its substrate, resorufin-beta-D-galactopyranoside, as the model system of the enzyme reaction. With a single experiment on the chip, we determined the key parameters for the enzyme kinetics, K(m) and k(cat), and evaluated the effect of inhibitor concentrations on the reaction rates. This study provides a new tool for evaluating various effectors, such as inhibitors and cofactors, on the initial rate of an enzyme reaction, and it could be applied to a comprehensive bio/chemical reaction study.
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