In addition, atoms involved in the restraints must be sufficiently near one another, often less than half the shortest box edge away, to avoid problems due to periodic boundary conditions and the minimum-image convention. While larger separations would not in principle be a problem, restraints in certain simulation packages like GROMACS do not smoothly handle the minimum-image convention. For example, in one system (estrogen receptor alpha), a small movement of atoms involved in the restraints led to one of the restrained dihedral angles jumping between being computed "through" versus "around" the box, based on the minimum-image convention. The sudden jumps in dihedral angle due to these imaging issues then resulted in the decoupled ligand, even though restrained, leaving the binding site. Of course, this was simply an artifact of periodicity -but because of the handling of periodicity in the calculation of restraints in GROMACS, this resulted in sudden jumps in restraint energy/forces and caused problems. Thus, we adjusted our restraints selection procedure to avoid this problem.