Pre-steady-state kinetics for hydrolysis of insoluble cellulose by cellobiohydrolase Cel7A.

ABSTRACT: The transient kinetic behavior of enzyme reactions prior to the establishment of steady state is a major source of mechanistic information, yet this approach has not been utilized for cellulases acting on their natural substrate, insoluble cellulose. Here, we elucidate the pre-steady-state regime for the exo-acting cellulase Cel7A using amperometric biosensors and an explicit model for processive hydrolysis of cellulose. This analysis allows the identification of a pseudo-steady-state period and quantification of a processivity number as well as rate constants for the formation of a threaded enzyme complex, processive hydrolysis, and dissociation, respectively. These kinetic parameters elucidate limiting factors in the cellulolytic process. We concluded, for example, that Cel7A cleaves abo

SUBMITTER: Cruys-Bagger N 

PROVIDER: S-EPMC3365755 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature