Open AccessProbing the role of cation-π interaction in the thermotolerance and catalytic performance of endo-polygalacturonases
Author(s) -
Tao Tu,
Yeqing Li,
Xiaoyun Su,
Kun Meng,
Rui Ma,
Yuan Wang,
Bin Yao,
Zhemin Lin,
Huiying Luo
Publication year - 2016
Publication title -
scientific reports
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.24
H-Index - 213
ISSN - 2045-2322
DOI - 10.1038/srep38413
Subject(s) - thermostability , pectinase , chemistry , mutant , hydrolysis , mutagenesis , bioconversion , catalysis , enzyme kinetics , substrate (aquarium) , catalytic efficiency , biochemistry , enzyme , site directed mutagenesis , glycoside hydrolase , stereochemistry , active site , biology , gene , fermentation , ecology
Understanding the dynamics of the key pectinase, polygalacturonase, and improving its thermotolerance and catalytic efficiency are of importance for the cost-competitive bioconversion of pectic materials. By combining structure analysis and molecular dynamics (MD) simulations, eight mutagenesis sites having the potential to form cation-π interactions were identified in the widely used fungal endo-polygalacturonase PG63. In comparison to the wild-type, three single mutants H58Y, T71Y and T304Y showed improved thermostability (the apparent T m s increased by 0.6−3.9 °C) and catalytic efficiency (by up to 32-fold). Chromatogram analysis of the hydrolysis products indicated that a larger amount of shorter sugars were released from the polygalacturonic acid by these three mutants than by the wild-type. MD analysis of the enzyme-substrate complexes illustrated that the mutants with introduced cation-π interaction have modified conformations of catalytic crevice, which provide an enviable environment for the catalytic process. Moreover, the lower plasticity of T3 loop 2 at the edge of the subsite tunnel appears to recruit the reducing ends of oligogalacturonide into the active site tunnel and initiates new hydrolysis reactions. This study demonstrates the importance of cation-π interaction in protein conformation and provides a realistic strategy to enhance the thermotolerance and catalytic performance of endo-polygalacturonases.