Kinetic Study on Enzymatic Hydrolysis of Cellulose in an Open, Inhibition-Free System.

Due to the complexity of cellulases and the requirement of enzyme adsorption on cellulose prior to reactions, it is difficult to evaluate their reaction with a general mechanistic scheme. Nevertheless, it is of great interest to come up with an approximate analytic description of a valid model for the purpose of developing an intuitive understanding of these complex enzyme systems. Herein, we used the surface plasmonic resonance method to monitor the action of a cellobiohydrolase by itself, as well as its mixture with a synergetic endoglucanase, on the surface of a regenerated model cellulose film, under continuous flow conditions. We found a phenomenological approach by taking advantage of the long steady state of cellulose hydrolysis in the open, inhibition-free system. This provided a direct and reliable way to analyze the adsorption and reaction processes with a minimum number of fitting parameters. We investigated a generalized Langmuir-Michaelis-Menten model to describe a full set of kinetic results across a range of enzyme concentrations, compositions, and temperatures. The overall form of the equations describing the pseudo-steady-state kinetics of the flow-system shares some interesting similarities with the Michaelis-Menten equation. The use of familiar Michaelis-Menten parameters in the analysis provides a unifying framework to study cellulase kinetics. The strategy may provide a shortcut for approaching a quantitative while intuitive understanding of enzymatic degradation of cellulose from top to bottom. The open system approach and the kinetic analysis should be applicable to a variety of cellulases and reaction systems to accelerate the progress in the field.