Effects of Site-Directed Mutations on the Communicability between Local Segments and Binding Pocket Distortion of Engineered GH11 Xylanases Visualized through Network Topology Analysis

Abstract

Mutations occurred within the binding pocket of enzymes directly modified the interaction network between an enzyme and its substrate. However, some mutations affecting the catalytic efficiency occurred far from the binding pocket and the explanation regarding mechanisms underlying the transmission of the mechanical signal from the mutated site to the binding pocket was lacking. In this study, network topology analysis was used to characterize and visualize the changes of interaction networks caused by site-directed mutations on a GH11 xylanase from our previous study. For each structure, coordinates from molecular dynamics (MD) trajectory were obtained to create networks of representative atoms from all protein and xylooligosaccharide substrate residues, in which edges were defined between pairs of residues within a cutoff distance. Then, communicability matrices were extracted from the network to provide information on the mechanical signal transmission from the number of possible paths between any residue pairs or local protein segments. The analysis of subgraph centrality and communicability clearly showed that site-direct mutagenesis at non-reducing or reducing ends caused binding pocket distortion close to the opposite ends and created denser interaction networks. However, site-direct mutagenesis at both ends cancelled the binding pocket distortion, while enhancing the thermostability. Therefore, the network topology analysis tool on the atomistic simulations of engineered proteins could play some roles in protein design for the minimization to the correction of binding pocket tilting, which could affect the functionality and efficacy of enzymes.