Synergistic effects between the additions of a disulphide bridge and an N-terminal hydrophobic sidechain on the binding pocket tilting and enhanced Xyn11A activity.

Synergistic effects between the additions of a disulphide bridge and an N-terminal hydrophobic sidechain on the binding pocket tilting and enhanced Xyn11A activity.
Arch Biochem Biophys. 2019 Aug 08;:108068
Authors: Boonyaputthikul H, Muhammad A, Roekring S, Rattanarojpong T, Khunrae P, Sutthibutpong T
Abstract
Synergistic effect of distal site-directed mutations and molecular mechanisms on the enhanced thermostability of GH11 xylanase from B. firmus Strain K-1 (xyn11A) was investigated through enzyme activity assays and atomistic molecular dynamics (MD) simulation. From the experiment, single N-terminal leucine substitution at K40L caused a significant drop in enzymatic activity. However, the addition of a disulphide bond at S100C/N147C, along with the K40L mutation enhanced the enzymatic activity at room temperature. Molecular mechanisms on the improvement of enzymatic activity were addressed through atomistic molecular dynamics (MD) simulations of enzyme-substrate complexes. Conformational analysis of the right-hand-shaped GH11 protein structures showed that K40L mutation 'tilted' the Palm region away from the Pinky finger at N-terminus and S100C/N147C tilted the Palm region towards the Pinky finger at N-terminus, which destabilized the binding complexes. The extended hydrophobic cluster formed within the K40L/S100C/N147C mutant stabilized the loops associated with the N-terminus and the Thumb region, which facilitated substrate b...

Source: Archives of Biochemistry and Biophysics - August 7, 2019 Category: Biochemistry Authors: Boonyaputthikul H, Muhammad A, Roekring S, Rattanarojpong T, Khunrae P, Sutthibutpong T Tags: Arch Biochem Biophys Source Type: research