I am using the FastDesign protocol in RosettaScripts to design some residues in the neighborhood of a ligand, specifically with constraints of N-O pair distance for NAD binding.
I've added constraints for the two N-O pairs to be 3.0 and 3.3 angstroms each with harmonic function and sd of 0.5, but somehow the distances are not kept below ~4.0 at all. (I've just observed a structure with over 6.0 angstrom distances for the two pairs.
I'm using talaris2014 score function and added a line in the xml file to re-weight atom_pair_constraint with a value of 1.0.
Is there something obvious that I'm missing?