Short-time dynamics of pH-dependent conformation and substrate binding in the active site of beta-glucosidases: A computational study.

Authors:
David F Flannelly
David F Flannelly
The Institute for Comparative and Environmental Toxicology
Thalia G Aoki
Thalia G Aoki
College of Agriculture and Life Sciences
College Station | United States
Dr. Ludmilla Aristilde, PhD
Dr. Ludmilla Aristilde, PhD
Cornell University
Associate Professor
Environmental Chemistry; Environmental Biochemistry; Environmental Engineering.
Ithaca, NY | United States

J Struct Biol 2015 Sep 6;191(3):352-64. Epub 2015 Jul 6.

The Institute for Comparative and Environmental Toxicology, College of Agricultural and Life Sciences, Cornell University, Ithaca, NY 14853, USA; Department of Biological and Environmental Engineering, College of Agricultural and Life Sciences, Cornell University, Ithaca, NY 14853, USA. Electronic address:

The complete degradation of cellulose to glucose is essential to carbon turnover in terrestrial ecosystems and to engineered biofuel production. A rate-limiting step in this pathway is catalyzed by beta-glucosidase (BG) enzymes, which convert cellulobiose into two glucose molecules. The activity of these enzymes has been shown to vary with solution pH. However, it is not well understood how pH influences the enzyme conformation required for catalytic action on the substrate. A structural understanding of this pH effect is important for predicting shifts in BG activity in bioreactors and environmental matrices, in addition to informing targeted protein engineering. Here we applied molecular dynamics simulations to explore conformational and substrate binding dynamics in two well-characterized BGs of bacterial (Clostridium cellulovorans) and fungal (Trichoderma reesei) origins as a function of pH. The enzymes were simulated in an explicit solvated environment, with NaCl as electrolytes, at their prominent ionization states obtained at pH 5, 6, 7, and 7.5. Our findings indicated that pH-dependent changes in the ionization states of non-catalytic residues localized outside of the immediate active site led to pH-dependent disruption of the active site conformation. This disruption interferes with favorable H-bonding interactions with catalytic residues required to initiate catalysis on the substrate. We also identified specific non-catalytic residues that are involved in stabilizing the substrate at the optimal pH for enzyme activity. The simulations further revealed the dynamics of water-bridging interactions both outside and inside the substrate binding cleft during structural changes in the enzyme-substrate complex. These findings provide new structural insights into the pH-dependent substrate binding specificity in BGs.

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http://dx.doi.org/10.1016/j.jsb.2015.07.002DOI Listing
September 2015
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