Characterization and Identification of The Key Residues Abound The Activity Site Tunnel of Trehalose Synthase from Pseudomonas Stutzeri Qlu3

摘要

The conversion of maltose into trehalose has important applications in the manufacture of food and other products. The enzyme trehalose synthase (TreS) catalyzes the interconversion of maltose and trehalose with glucose as a byproduct. In this study, treS was cloned from Pseudomonas stutzeri Qlu3 genomic DNA. We predicted the structural characteristics of recombinant TreS bound to its substrate by using homology modeling and flexible docking studies of the enzyme-substrate system. These analyses showed six amino acids (Phell5, Phe255, Arg292, Asp403, Asp294, and Glu338) that interact extensively with the substrate during catalysis. In addition, an enclosed active site tunnel was revealed that controls substrate movement during intramolecular isomerization. Disruption of the tunnel by removing two loops led to total loss of isomerization activity. The A309E mutant showed increased isomerase activity and decreased hydrolase activity. In contrast, the Q219R, T308E, and L341Q mutants showed decreased isomerase activity and increased hydrolase activity. These results suggest that the size of the tunnel can influence isomerase activity and hydrolase activity. Therefore, our results exhibited the TreS from Pseudomonas stutzeri Qlu3 have potential industrial use and the analysis in the structure has additional aid for the further molecular modification.