Modeling the activity burst in the initial phase of cellulose hydrolysis by the processive cellobiohydrolase Cel7A

Abstract

The hydrolysis of cellulose by processive cellulases, such as exocellulase Tr Cel7A from Trichoderma reesei , is typically characterized by an initial burst of high activity followed by a slowdown, often leading to incomplete hydrolysis of the substrate. The origins of these limitations to cellulose hydrolysis are not yet fully understood. Here, we propose a new model for the initial phase of cellulose hydrolysis by processive cellulases, incorporating a bound but inactive enzyme state. The model, based on ordinary differential equations, accurately reproduces the activity burst and the subsequent slowdown of the cellulose hydrolysis and describes the experimental data equally well or better than the previously suggested model. We also derive steady‐state expressions that can be used to describe the pseudo‐steady state reached after the initial activity burst. Importantly, we show that the new model predicts the existence of an optimal enzyme‐substrate affinity at which the pseudo‐steady state hydrolysis rate is maximized. The model further allows the calculation of glucose production rate from the first cut in the processive run and reproduces the second activity burst commonly observed upon new enzyme addition. These results are expected to be applicable also to other processive enzymes.

Larger Intrinsic Rate Constants of Alpha-amylase is Possible if Intrinsic Forward Rate Constant is ≠ Diffusion limited Rate of Encounter

“…This research has given quantitative effect to theoretical input on the problem of intrinsic rate constants. There is a need, however, to add that work is going on in the area of the kinetics of cellulose hydrolysis by cellulases where, in one instance, the apparent processivity was observed to be typically smaller than the intrinsic processivity defined on the basis of a theoretical model using apparent rate constants [21]; this is quite similar to the procedure in this research. In order to understand the role of the catalytic site, the catalytic domain has been modeled as a one-dimensional stochastic "walker" that may only step in the forward direction as governed by an intrinsic rate constant, [22].…”

Model of Processive Catalysis with Site Clustering and Blocking and Its Application to Cellulose Hydrolysis

“…The results of the simulations are also in agreement with a simple analytical three-state model, which assumes the existence of an active, hydrolyzing bound state and an abstract inactive bound state. 45 The analytical model described the experimental data best when assuming that the initially free particle first binds to the active state and later enters the inactive state. The CB model introduced here offers a concrete picture as to the nature of the active and inactive states.…”

Directly and Indirectly Determinable Rate Constants in Michaelian Enzyme-Catalyzed Reactions

“…On a matter of general interest and importance arising from this study in light of the concern for validity, stability of ES, etc., one begins by stating that the vast array of literature materials covering enzyme kinetics is evidence of the importance of kinetic parameters; besides, there are no kinetic studies without rates and cognate rate constants. So, a lot of research papers on the kinetics of enzyme-catalyzed reactions with associated models abound in the literature [29][30][31][32][33][34]. Also, researchers [35,36] have published kinetic equations of enzyme-catalyzed reactions using kinetic schemes.…”