Abstract
This paper reports a method to characterize the kinetic constants for the action of enzymes on immobilized substrates. This example uses cutinase, a serine esterase that hydrolyzes 4-hydroxyphenyl valerate moieties that are immobilized on a self-assembled monolayer of alkanethiolates on gold. The product of the enzyme reaction is a hydroquinone, which is redox active and therefore permits the use of cyclic voltammetry to monitor the extent of reaction in situ. A kinetic model based on the Michaelis-Menten formalism is used to analyze the dependence of initial rates of reaction on both the substrate density and the enzyme concentration. The resulting value of k cat/KM for the interfacial reaction is comparable to that for a homogeneous phase reaction with a substrate of similar structure. This strategy of using monolayers presenting substrates for the enzyme and cyclic voltammetry to measure reaction rates provides quantitative and real-time information on reaction rates and permits a level of analysis of interfacial enzyme reactions that to date has been difficult to realize.
Original language | English (US) |
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Pages (from-to) | 5578-5583 |
Number of pages | 6 |
Journal | Langmuir |
Volume | 23 |
Issue number | 10 |
DOIs | |
State | Published - May 8 2007 |
ASJC Scopus subject areas
- General Materials Science
- Condensed Matter Physics
- Surfaces and Interfaces
- Spectroscopy
- Electrochemistry