Homology modeling and molecular dynamics (MD) simulations
The 3D structural of Pc APSK and its mutants were constructed based on its X‐ray crystal structures (PDB ID: 1M7H) (Lansdon et al., 2002). The 3D structure of ADP was obtained from the ChemSpider Ultra 11.0 program (http://www.chemspider.com/ ). Nonpolar hydrogen atoms were added to the enzyme using the GROMACS 4.5.5 simulation package (http://www.gromacs.org/). The whole system was immersed in an explicit TIP 3P water box and extended with a thickness of at least 10 Å from the dissolved atoms in each dimension. MD simulations were performed with GROMACS 4.5.5 and an AMBER03 force field following the three main steps of energy minimization, system equilibration, and production protocols. The 50 ns MD simulations were performed using NAMD 2.12 (derived from “Not Another Molecular Dynamics program”, by University of Illinois) with the Charmm27 force field with a 2-fs time step at the temperature of 310 K. The enzyme and product were assigned to separate energy groups, and the binding energies between them were calculated as total intergroup potential energy. The MD simulations results were analyzed in Visual Molecular Dynamics software (VMD 1.9.3, by University of Illinois). All images of the structures shown here were generated using PyMol and CAVER. The two flexible regions (A and B) identified by MD simulations were as follows: region A comprised residues from L33 to V43, and B region included residues from Y137 to H181.