(608f) ORNL: Initial Binding of a Cellulose Chain Into the Cellulase Catalytic Tunnel

Cel7A proceeds along a single cellulose chain in a processive manner, cleaving one cellobiose unit per catalytic event. We have conducted extensive molecular dynamics (MD) simulations to understand the initial binding of a cellulose chain into the catalytic tunnel of the Trichoderma reesei Family 7 cellobiohydrolase (Cel7A). In unrestrained MD simulations, the cellulose chain diffuses into the tunnel by the length of a cellobiose unit. Among those residues lining the tunnel, the four tryptophan residues exhibit small structural fluctuations, and may thus play a role in guiding the substrate sliding, while the more dynamic polar residues are observed to promote the initial diffusion of the substrate into the tunnel. Further MD simulations show no evident preference for the reducing end or the non-reducing end binding of the cellulose chain, suggesting that the spontaneous initial binding is not driven by the dipole-dipole interaction between Cel7A and the cellulose chain. Rather, the shape complementarity of the cellulose chain end to the tunnel vestibule (involving local hydrophobic stacking and hydrogen bond interactions) plays a more dominant role. The potential of mean force (PMF) calculations, however, show a clear difference at -5 subsite between the reducing end and the non-reducing end binding, implicating that the selective hydrolysis of cellulose from the reducing end can be partially achieved through the preferential initial binding of cellulose to the Cel7A tunnel. The comparison of the PMFs for the wild type and the W40A mutant enzymes highlights the importance of the Trp40 residue in the initial threading of the cellulose chain to the catalytic tunnel.