CelS-Catalyzed Processive Cellulose Degradation and Cellobiose Extraction for the Production of Bioethanol

Bacterial cellulase enzymes are potent
candidates for the efficient
production of bioethanol, a promising alternative to fossil fuels,
from cellulosic biomass. These enzymes catalyze the breakdown of cellulose
in plant biomass into simple sugars and then to bioethanol. In the
absence of the enzyme, the cellulosic biomass is recalcitrant to decomposition
due to fermentation-resistant lignin and pectin coatings on the cellulose
surface, which make them inaccessible for hydrolysis. Cellobiohydrolase
CelS is a microbial enzyme that binds to cellulose fiber and efficiently
cleaves it into a simple sugar (cellobiose) by a repeated processive
chopping mechanism. The two contributing factors to the catalytic
reaction rate and the yield of cellobiose are the efficient product
expulsion from the product binding site of CelS and the movement of
the substrate or cellulose chain into the active site. Despite progress
in understanding product expulsion in other cellulases, much remains
to be understood about the molecular mechanism of processive action
of these enzymes. Here, nonequilibrium molecular dynamics simulations
using suitable reaction coordinates are carried out to investigate
the energetics and mechanism of the substrate dynamics and product
expulsion in CelS. The calculated free energy barrier for the product
expulsion is three times lower than that for the processive action
indicating that product removal is relatively easier and faster than
the sliding of the substrate to the catalytic active site. The water
traffic near the active site in response to the product expulsion
and the processive action is also explored.