Refinement without restraints generally led to significant deterioration of model qualities. The model quality changes without restraints in terms of Cα-RMSD, GDT-HA, lDDT, and SphereGrinder were -0.08 Å, -1.03, 0.0029, -1.12 at 298.15 K and 0.26 Å, -4.65, -0.0175, -3.53 at 360 K, respectively. These results confirm previous studies that found that restraints are necessary for successful refinement with the exception of a few successful cases involving very small protein models. 24