Immobilization of Cellulase Enzymes on Single-Walled Carbon Nanotubes for Recycling of Enzymes and Better Yield of Bioethanol Using Computer Simulations.

The utilization of microbial cellulase enzymes for transforming plant biomass into biofuel or bioethanol, which can serve as a substitute for fossil fuel, is a subject of growing interest. Nonetheless, large-scale production of biofuel using cellulases is not economically feasible as the extraction of these enzymes from diverse microorganisms is an expensive process. To address this issue, immobilizing the enzyme to a substrate material, e.g., carbon nanotubes (CNTs), to recycle without a significant decline in its catalytic activity is a promising solution. Due to the hydrophobic nature of CNTs, we employed molecular docking and network analysis methodologies to identify potential CNT-binding sites on the outer surface of a wild-type cellulase enzyme, CelS. Classical molecular dynamics simulations of CNT-bound CelS through one of the selected binding sites resulted in negligible changes in the secondary structure of the enzyme and its catalytic domain, implying the least possible effect on the catalytic activity post-immobilization. Furthermore, our study reveals that while the unfolding near the CNT-binding region in CelS is more pronounced when the enzyme is interacting with a wider CNT, resulting in enhanced contact area and improved binding affinity, its impact on the overall CelS structure is relatively less significant when compared to thinner CNTs. Particularly, CNTs of diameter ∼12 Å can serve as a favorable option for substrate materials in cellulase immobilization. Our study also provides critical insights into the binding mechanisms between cellulase and CNTs, which could lead to the development of more efficient biocatalysts for biofuel production.