2018 Denovo design tutorial 1 with PDB 2NNQ
Contents
2018 Denovo design with PDB 2NNQ (Focused)
Files Needed
Fragment Libraries
A focused fragment library will be used in this tutorial, in order to attempt building the same ligand. A focused fragment library can be generated using the same ligand.
Create a new directory for the fragment library.
mkdir fraglib
Inside the fraglib directory create a new input file for fragment generation.
touch fraglib.in
Generate the fragments by calling the input file through DOCK6
dock6 -i fraglib.in
Answer the prompted questions interactively using the following lines.
conformer_search_type flex write_fragment_libraries yes fragment_library_prefix fraglib fragment_library_freq_cutoff 1 fragment_library_sort_method freq fragment_library_trans_origin no use_internal_energy yes internal_energy_rep_exp 12 internal_energy_cutoff 100.0 ligand_atom_file /Path_to_file/2nnq_lig_withH.mol2 limit_max_ligands no skip_molecule no read_mol_solvation no calculate_rmsd no use_database_filter no orient_ligand yes automated_matching yes receptor_site_file /Path_to_file/selected_spheres.sph max_orientations 1000 critical_points no chemical_matching no use_ligand_spheres no bump_filter no score_molecules no atom_model all vdw_defn_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/vdw_AMBER_parm99.defn flex_defn_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/flex.defn flex_drive_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/flex_drive.tbl ligand_outfile_prefix output write_orientations no num_scored_conformers 1 rank_ligands no
Once fragment generation is completed, following files will be generated. (fraglib_linker.mol2, fraglib_rigid.mol2, fraglib_scaffold.mol2, fraglib_sidechain.mol2, fraglib_torenv.dat) Open the mol2 files using chimera and check if the fragments match with the ligand used for fragment generation.
Focused Denovo Growth
The generated fragments will be used to perform the de novo dock. Here the fragments will added together restricted by different properties like maximum layers, formal charge and molecular weight. Before each connection, DOCK will check with the torsion environment file created in frgment generation to see if that particular connection is seen before. If not the connection won't be made.
Create a new directory for denovo dock.
mkdir denovo
Create a new input file for denovo dock.
touch denovo.in
Run the input file through dock.
dock6 -i denovo.in
Answer the prompted question interactively using the following lines.
conformer_search_type flex write_fragment_libraries yes fragment_library_prefix fraglib fragment_library_freq_cutoff 1 fragment_library_sort_method freq fragment_library_trans_origin no use_internal_energy no ligand_atom_file ./candidate_mol_gen_frag.mol2 limit_max_ligands no skip_molecule no read_mol_solvation no calculate_rmsd no use_database_filter no orient_ligand no bump_filter no score_molecules no atom_model all vdw_defn_file ../../../dock/box/vdw_AMBER_parm99.defn flex_defn_file ../../../dock/dock/flex.defn flex_drive_file ../../../dock/dock/flex_drive.tbl ligand_outfile_prefix output write_orientations no num_scored_conformers 1 rank_ligands no
Focused Denovo Rescore
Generic de novo growth with PDB 2NNQ
In this section, the de novo growth will be done using a fragment library generated by many random ligands. This allows DOCK to generate novel ligands which are not restricted by the fragments and torsion environment of the crystal ligand.
In the previous section, single grid was used as the scoring function to generate the ligands. In this section, the interactions of the ligand with the most significant residues will be considered for the scoring function. In order to do so, multigrid scoring method where each of the significant residues will be described using unique grids, and a common grid for the rest of the residues.
Specifying Primary Residues
First, the significant residues for the binding of the ligand has to be specified. Create unique directory for the generic de novo growth. Create an input file. (rescore.in) Use the following lines to generate the input file.
conformer_search_type rigid use_internal_energy no ligand_atom_file ../2nnq_lig_withH.mol2 (use the ligand mol2 file) limit_max_ligands no skip_molecule no read_mol_solvation no calculate_rmsd no use_database_filter no orient_ligand no bump_filter no score_molecules yes contact_score_primary no contact_score_secondary no grid_score_primary no grid_score_secondary no multigrid_score_primary no multigrid_score_secondary no dock3.5_score_primary no dock3.5_score_secondary no continuous_score_primary no continuous_score_secondary no footprint_similarity_score_primary yes footprint_similarity_score_secondary no fps_score_use_footprint_reference_mol2 yes fps_score_footprint_reference_mol2_filename ../2nnq_lig_withH.mol2 (use the ligand mol2 file) fps_score_foot_compare_type Euclidean fps_score_normalize_foot no fps_score_foot_comp_all_residue no fps_score_choose_foot_range_type threshold fps_score_vdw_threshold 1 fps_score_es_threshold 0.5 fps_score_hb_threshold 0.5 fps_score_use_remainder yes fps_score_receptor_filename ../2nnq_rec.mol2 (use the mol2 file for the receptor) fps_score_vdw_att_exp 6 fps_score_vdw_rep_exp 12 fps_score_vdw_rep_rad_scale 1 fps_score_use_distance_dependent_dielectric yes fps_score_dielectric 4.0 fps_score_vdw_fp_scale 1 fps_score_es_fp_scale 1 fps_score_hb_fp_scale 0 pharmacophore_score_secondary no descriptor_score_secondary no gbsa_zou_score_secondary no gbsa_hawkins_score_secondary no SASA_score_secondary no amber_score_secondary no minimize_ligand no atom_model all vdw_defn_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/vdw_AMBER_parm99.defn flex_defn_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/flex.defn flex_drive_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/flex_drive.tbl ligand_outfile_prefix rescore.out write_footprints yes write_hbonds no write_orientations no num_scored_conformers 1 rank_ligands no
This will generate three output files.
2nnq.footprint_rescore.out rescore.out_footprint_scored.txt rescore.out_scored.mol2
Use these files to specify the primary residues. Create a new file specify.sh using the following lines which are taken from previous tutorials and altered accordingly.
#!/bin/bash
grep -A 1 "range_union" footprint.out |
grep -v "range_union" |
grep -v "\-" |
sed -e '{s/,/\n/g}' |
sed -e '{s/ //g}' |
sed '/^$/d' |
sort -n |
uniq > temp.dat
for i in `cat temp.dat`; do printf "%0*d\n" 3 $i; done > 2nnq.primary_residues.dat
for RES in `cat temp.dat`
do
grep " ${RES} " rescore.out_footprint_scored.txt |
awk -v temp=${RES} '{if ($2 == temp) print $0;}' |
awk '{print $1 " " $3 " " $4}' >> reference.txt
done
grep "remainder" rescore.out_footprint_scored.txt |
sed -e '{s/,/ /g}' |
tr -d '\n' |
awk '{print $2 " " $3 " " $6}' >> reference.txt
mv reference.txt 2nnq.reference.txt
rm temp.dat
export AMBERHOME="/gpfs/projects/AMS536/zzz.programs/amber16"
Use either of the following commands to specify the primary residues using the above file.
chmod 770 specify.sh ./specify.sh
or
bash specify.sh
This will generate another two files which specify the primary residues of the binding site of your receptor.
2nnq.primary_residues.dat 2nnq.reference.txt
If it is successful, 25 residues should be specified as primary residues. (this is just for this system and the number of primary residues can change from system to system)
In the next section, grids will be generated for the primary residues and a common grid for the rest of the residues.
Generation of grids
Create a new directory (multigrid) This will be done by a script file and two input files. The structure of each file is given below.
First input file (2nnq.multigrid.in)
compute_grids yes grid_spacing 0.4 output_molecule yes contact_score no chemical_score no energy_score yes energy_cutoff_distance 9999 atom_model a attractive_exponent 6 repulsive_exponent 9 distance_dielectric yes dielectric_factor 4 bump_filter yes bump_overlap 0.75 receptor_file temp.mol2 box_file ../../../dock/3.boxgrid/2nnq.box.pdb (use the virtual box generated in dock previously) vdw_definition_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/vdw_AMBER_parm99.defn chemical_definition_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/chem.defn score_grid_prefix temp.rec receptor_out_file temp.rec.grid.mol2
Second input file (2nnq.reference_multigrid.in)
conformer_search_type rigid use_internal_energy yes internal_energy_rep_exp 12 internal_energy_cutoff 100.0 ligand_atom_file ../../2nnq_lig_withH.mol2 limit_max_ligands no skip_molecule no read_mol_solvation no calculate_rmsd no use_database_filter no orient_ligand no bump_filter no score_molecules yes contact_score_primary no contact_score_secondary no grid_score_primary no grid_score_secondary no multigrid_score_primary yes multigrid_score_secondary no multigrid_score_rep_rad_scale 1.0 multigrid_score_vdw_scale 1.0 multigrid_score_es_scale 1.0 multigrid_score_number_of_grids 26 (Should be the total number of primary residues + 1) multigrid_score_grid_prefix0 2nnq.resid_014 multigrid_score_grid_prefix1 2nnq.resid_016 multigrid_score_grid_prefix2 2nnq.resid_019 multigrid_score_grid_prefix3 2nnq.resid_020 multigrid_score_grid_prefix4 2nnq.resid_036 multigrid_score_grid_prefix5 2nnq.resid_037 multigrid_score_grid_prefix6 2nnq.resid_038 multigrid_score_grid_prefix7 2nnq.resid_040 multigrid_score_grid_prefix8 2nnq.resid_051 multigrid_score_grid_prefix9 2nnq.resid_057 multigrid_score_grid_prefix10 2nnq.resid_058 multigrid_score_grid_prefix11 2nnq.resid_060 multigrid_score_grid_prefix12 2nnq.resid_072 multigrid_score_grid_prefix13 2nnq.resid_074 multigrid_score_grid_prefix14 2nnq.resid_075 multigrid_score_grid_prefix15 2nnq.resid_076 multigrid_score_grid_prefix16 2nnq.resid_078 multigrid_score_grid_prefix17 2nnq.resid_093 multigrid_score_grid_prefix18 2nnq.resid_104 multigrid_score_grid_prefix19 2nnq.resid_106 multigrid_score_grid_prefix20 2nnq.resid_115 multigrid_score_grid_prefix21 2nnq.resid_116 multigrid_score_grid_prefix22 2nnq.resid_117 multigrid_score_grid_prefix23 2nnq.resid_126 multigrid_score_grid_prefix24 2nnq.resid_128 multigrid_score_grid_prefix25 2nnq.resid_remaining (rest of the residues) multigrid_score_fp_ref_mol no multigrid_score_fp_ref_text yes multigrid_score_footprint_text ../2nnq.reference.txt (generated earlier when specifying the primary residues) multigrid_score_foot_compare_type Euclidean multigrid_score_normalize_foot no multigrid_score_vdw_euc_scale 1.0 multigrid_score_es_euc_scale 1.0 dock3.5_score_secondary no continuous_score_secondary no footprint_similarity_score_secondary no pharmacophore_score_secondary no descriptor_score_secondary no gbsa_zou_score_secondary no gbsa_hawkins_score_secondary no SASA_score_secondary no amber_score_secondary no minimize_ligand yes simplex_max_iterations 1000 simplex_tors_premin_iterations 0 simplex_max_cycles 1 simplex_score_converge 0.1 simplex_cycle_converge 1.0 simplex_trans_step 1.0 simplex_rot_step 0.1 simplex_tors_step 10.0 simplex_random_seed 0 simplex_restraint_min no atom_model all vdw_defn_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/vdw_AMBER_parm99.defn flex_defn_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/flex.defn flex_drive_file /gpfs/projects/AMS536/zzz.programs/dock6/parameters/flex_drive.tbl ligand_outfile_prefix output write_footprints no write_orientations no num_scored_conformers 1 rank_ligands no
Use the following script to generate the grids. (multigrid.sh)
export PRIMARY_RES=` cat ../2nnq.primary_residues.dat | sed -e 's/\n/ /g' `
export DOCKHOME="/gpfs/projects/AMS536/zzz.programs/dock6_new"
python ${DOCKHOME}/bin/multigrid_fp_gen.py ../../2nnq_rec.mol2 2nnq.resid 2nnq.multigrid.in ${PRIMARY_RES}
rm temp.mol2
rm 2nnq.resid_*.rec.grid.mol2
${DOCKHOME}/bin/dock6 -i 2nnq.reference_multigridmin.in -o 2nnq.reference_multigridmin.out
mv output_scored.mol2 2nnq.lig.multigridmin.mol2
cp 2nnq.lig.multigridmin.mol2 ../multi-grid
