MD studies.

In order to gain insights into the intrinsic conformational properties of the cyclic peptide, a molecular dynamics (MD) simulation was conducted using the structure of the peptide detected in the complex with Fab C2 as the starting model. The molecular dynamics simulation was performed using the GROMACS software package (version 4.5.5) (47), the AMBER99sb force field, and TIP4P as the water model. The peptide was immersed in a cubic box of 4.50 by 4.50 by 4.50 nm³ containing 2,933 water molecules. The simulation was run with periodic boundary conditions. The temperature and pressure of the systems were stabilized at 300 K and 1 atm, respectively. Energies were minimized by fixing the protein atoms, and then the simulation was run without restraints. The time scale of the simulation was 250 ns with a time step of 0.002 ps. The particle mesh Ewald (PME) method (grid spacing, 0.12 nm) was used to calculate the electrostatic interactions. A cutoff of 10 Å was applied to treat Lennard-Jones interactions. Bond lengths were constrained using the Linear Constraint Solver (LINCS) algorithm. Trajectories were checked to assess the quality of the simulation using GROMACS routines. H-bond interactions were identified on the basis of the cutoffs for the hydrogen donor-acceptor angle (∼30°) and the donor-acceptor distance (∼3.5 Å) by using GROMACS utilities (47).