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Published August 2012 | public
Journal Article

Computational Thermostable Cellulase Engineering for Enhanced Biofuel Production


Generating biofuels from cellulosic material may mitigate the detrimental environmental effects of current production schemes. Common agricultural byproducts and algal feedstocks are cellulosic in nature, providing more biomass than grain sources while minimizing agricultural intensification. Employing cellulosic feedstocks, however, requires the use of relatively expensive cellulases, enzymes capable of cleaving cellulose into fermentable sugars. One method of reducing cellulase cost relies on an idea encapsulated in the Arrhenius equation, that reaction rates increase as temperature increases. Employing thermostable cellulases would not only minimize process time, but would also reduce the amount of enzyme necessary per gallon of synthesized fuel. We are currently computationally engineering hyperthermostable variants of Cel5A from the fungus Hypocrea jecorina (Hj_ Cel5A), one of the most heavily employed cellulases in the biofuels industry. To obtain a starting scaffold for design purposes, we have recently solved the structure of this enzyme. When compared to a thermostable homolog, the Hj_Cel5A structure demonstrates long, solvent-exposed loops. As such, we are combining traditional core-repacking algorithms with loop truncation software to generate thermostable variants. To ensure loop truncation does not disrupt protein stability, we utilize an algorithm to predict loop trajectories around the mutated sites, and then perform sequence optimization on the new structure. Here we present the crystal structure of H. jecorina Cel5A to 2.1 Å, data demonstrating the robustness of our design methodology, and preliminary designs that await biochemical testing.

Additional Information

This work was made possible through funding from the National Science Foundation.

Additional details

August 19, 2023
October 18, 2023