Cellulose Binding in the Processive Cellulase Cel7A

Cellobiohydrolases effectively degrade cellulose and are of biotechnological interest because they can convert lignocellulosic biomass to fermentable sugars. Using a fluorescence-based method for real-time measurements of complexation and decomplexation, the authors followed the processive cellulase Cel7A and its insoluble substrate, cellulose. Wild type Cel7A and several variants in which one or two of four highly conserved Trp residues in the binding tunnel had been replaced with Ala, were investigated.

Panel A shows a simplified illustration of the reaction mechanism of Cel7A. Panel B : The enlargement shows the structure of the bound cello-nonaose ligand (green/red sticks) and the location of the four conserved Trp residues. Numbers in the enlargement identify the subsites ; the scissile bond is between subsites +1 and -1. PDB 4C4C.

The results of this analysis suggested a transition state for complexation and decomplexation in which the reducing end of the ligand is close to the tunnel entrance, whereas the rest of the binding tunnel is empty. This structure defines the highest free-energy barrier of the overall catalytic cycle and hence governs the turnover rate of this industrially important enzyme.