The sequence_tolerance application is using the standard Rosetta file reading, so it should be sufficient to apply the params file like you would any other Rosetta application — that is, pass it in with the “-extra_res_fa” flag.
I haven’t tried it, but I think this should work with the sequence_tolerance application. The only caveat is that sequence_tolerance will probably not like trying to “design” your ligand, so you’ll want to provide a resfile which turns off design to the ligand.
You only need separate params files for ligands which are distinct chemical entities. Different conformations of the same ligand (e.g. formed by rotating around internal bonds) can be handled with the same params file.
Generally, though, what people do is create multiple conformers of the ligand in a multi-entry mol/sdf/mol2 file, and then give that to molfile_to_params.py – this allows you to specify a conformers file, and Rosetta will use the multiple conformers when packing – exactly like it does with the multiple rotamers of the protein side chains.
If you’re setting the ligand not to repack, though, a single params file should work for multiple conformers.