Parameter determination and validation for a mechanistic model of the enzymatic saccharification of cellulose-Iß.

Cost-effective production of fuels and chemicals from lignocellulosic biomass often involves enzymatic saccharification, which has been the subject of intense research and development. Recently, a mechanistic model for the enzymatic saccharification of cellulose has been developed that accounts for distribution of cellulose chain lengths, the accessibility of insoluble cellulose to enzymes, and the distinct modes of action of the component cellulases [Griggs et al. (2012) Biotechnol. Bioeng., 109(3):665-675; Griggs et al. (2012) Biotechnol. Bioeng., 109(3):676-685]. However, determining appropriate values for the adsorption, inhibition, and rate parameters required further experimental investigation. In this work, we performed several sets of experiments to aid in parameter estimation and to quantitatively validate the model. Cellulosic materials differing in degrees of polymerization and crystallinity (α-cellulose-Iß and highly crystalline cellulose-Iß ) were digested by component enzymes (EGI/CBHI/ßG) and by mixtures of these enzymes. Based on information from the literature and the results from these experiments, a single set of model parameters was determined, and the model simulation results using this set of parameters were compared with the experimental data of total glucan conversion, chain-length distribution, and crystallinity. Model simulations show significant agreement with the experimentally derived glucan conversion and chain-length distribution curves and provide interesting insights into multiple complex and interacting physico-chemical phenomena involved in enzymatic hydrolysis, including enzyme synergism, substrate accessibility, cellulose chain length distribution and crystallinity, and inhibition of cellulases by soluble sugars.

A mechanistic model for enzymatic saccharification of cellulose using continuous distribution kinetics II: cooperative enzyme action, solution kinetics, and product inhibition.

The projected cost for the enzymatic hydrolysis of cellulosic biomass continues to be a barrier for the commercial production of liquid transportation fuels from renewable feedstocks. Predictive models for the kinetics of the enzymatic reactions will enable an improved understanding of current limitations, such as the slow-down of the overall conversion rate, and may point the way for more efficient utilization of the enzymes in order to achieve higher conversion yields. A mechanistically based kinetic model for the enzymatic hydrolysis of cellulose was recently reported in Griggs et al. (2011) (Part I). In this article (Part II), the enzyme system is expanded to include solution-phase kinetics, particularly cellobiose-to-glucose conversion by ß-glucosidase (ßG), and novel adsorption and product inhibition schemes have been incorporated, based on current structural knowledge of the component enzymes. Model results show cases of cooperative and non-cooperative hydrolysis for an enzyme system consisting of EG(I) and CBH(I). The model is used to explore various potential rate-limiting phenomena, such as substrate accessibility, product inhibition, sterically hindered enzyme adsorption, and the molecular weight of the cellulose substrate.

A mechanistic model for enzymatic saccharification of cellulose using continuous distribution kinetics I: depolymerization by EGI and CBHI.

A mechanistically based kinetic model for the enzymatic hydrolysis of cellulosic biomass has been developed that incorporates the distinct modes of action of cellulases on insoluble cellulose polymer chains. Cellulose depolymerization by an endoglucanase (endoglucanase I, EG(I) ) and an exoglucanase (cellobiohydrolase I, CBH(I)) is modeled using population-balance equations, which provide a kinetic description of the evolution of a polydisperse distribution of chain lengths. The cellulose substrate is assumed to have enzyme-accessible chains and inaccessible interior chains. EG(I) is assumed to randomly cleave insoluble cellulose chains. For CBH(I), distinct steps for adsorption, complexation, processive hydrolysis, and desorption are included in the mechanistic description. Population-balance models that employ continuous distributions track the evolution of the spectrum of chain lengths, and do not require solving equations for all chemical species present in the reacting mixture, resulting in computationally efficient simulations. The theoretical and mathematical development needed to describe the hydrolysis of insoluble cellulose chains embedded in a solid particle by EG(I) and CBH(I) is given in this article (Part I). Results for the time evolution of the distribution of chain sizes are provided for independent and combined enzyme hydrolysis. A companion article (Part II) incorporates this modeling framework to study cellulose conversion processes, specifically, solution kinetics, enzyme inhibition, and cooperative enzymatic action.