View source for 2019 Denovo design tutorial 3 with PDB 3JQZ

This tutorial is a continuation of the [[2019 DOCK tutorial 3 with PDBID 3JQZ|virtual screen tutorial for 3JQZ]], with the same directory and naming structure.

Below is a figure showing a ligand generated by Denovo growth from a generic oxygen fragment (magenta) compared to the crystal lignad (blue). The chosen molecule was selected based on the highest volume overlap score with the crystal ligand. 

[[File:Denovo oxygen anchor 1721.jpg|thumb|center|1000px|Denovo growth using a generic oxygen anchor (magenta) compared to the crystal ligand (blue)]]


=='''I.'' de-novo'' design with PDB 3JQZ (Focused)'''==

===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                                             ../001.dockprep/3jqz_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                                           ../002.surface_spheres/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.

NEEDS TO BE FIXED - John Bickel - 3/11/19

 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                                                /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

===Focused Denovo Rescore===

Create a new directory
 mkdir focuseddenovorescore

Copy your reference .mol2 file into this directory

Create an input file
 touch rescore.in

Complete the input file using the automated questions from dock
 dock6 -i rescore.in

 conformer_search_type                                        rigid
 use_internal_energy                                          yes
 internal_energy_rep_exp                                      12
 internal_energy_cutoff                                       100.0
 ligand_atom_file                                             output.denovo_build.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                                      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                           no
 footprint_similarity_score_secondary                         no
 pharmacophore_score_primary                                  no
 pharmacophore_score_secondary                                no
 descriptor_score_primary                                     yes
 descriptor_score_secondary                                   no
 descriptor_use_grid_score                                    no
 descriptor_use_multigrid_score                               no
 descriptor_use_continuous_score                              no
 descriptor_use_footprint_similarity                          yes
 descriptor_use_pharmacophore_score                           yes
 descriptor_use_tanimoto                                      yes
 descriptor_use_hungarian                                     yes
 descriptor_use_volume_overlap                                yes
 descriptor_fps_score_use_footprint_reference_mol2            yes
 descriptor_fps_score_footprint_reference_mol2_filename       candidate_mol_gen_frag.mol2
 descriptor_fps_score_foot_compare_type                       Euclidean
 descriptor_fps_score_normalize_foot                          no
 descriptor_fps_score_foot_comp_all_residue                   yes
 descriptor_fps_score_receptor_filename                       ../001.dockprep/3jqz.rec.withH.charged.mol2
 descriptor_fps_score_vdw_att_exp                             6
 descriptor_fps_score_vdw_rep_exp                             12
 descriptor_fps_score_vdw_rep_rad_scale                       1
 descriptor_fps_score_use_distance_dependent_dielectric       yes
 descriptor_fps_score_dielectric                              4.0
 descriptor_fps_score_vdw_fp_scale                            1
 descriptor_fps_score_es_fp_scale                             1
 descriptor_fps_score_hb_fp_scale                             0
 descriptor_fms_score_use_ref_mol2                            yes
 descriptor_fms_score_ref_mol2_filename                       candidate_mol_gen_frag.mol2
 descriptor_fms_score_write_reference_pharmacophore_mol2      no
 descriptor_fms_score_write_reference_pharmacophore_txt       no
 descriptor_fms_score_write_candidate_pharmacophore           no
 descriptor_fms_score_write_matched_pharmacophore             no
 descriptor_fms_score_compare_type                            overlap
 descriptor_fms_score_full_match                              yes
 descriptor_fms_score_match_rate_weight                       5.0
 descriptor_fms_score_match_dist_cutoff                       1.0
 descriptor_fms_score_match_proj_cutoff                       0.7071
 descriptor_fms_score_max_score                               20
 descriptor_fingerprint_ref_filename                          candidate_mol_gen_frag.mol2
 descriptor_hms_score_ref_filename                            candidate_mol_gen_frag.mol2
 descriptor_hms_score_matching_coeff                          -5
 descriptor_hms_score_rmsd_coeff                              1
 descriptor_volume_score_reference_mol2_filename              candidate_mol_gen_frag.mol2
 descriptor_volume_score_overlap_compute_method               analytical
 descriptor_weight_fps_score                                  1
 descriptor_weight_pharmacophore_score                        1
 descriptor_weight_fingerprint_tanimoto                       -1
 descriptor_weight_hms_score                                  1
 descriptor_weight_volume_overlap_score                       -1
 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
 chem_defn_file                                               /gpfs/projects/AMS536/zzz.programs/dock6/parameters/chem.defn
 pharmacophore_defn_file                                      /gpfs/projects/AMS536/zzz.programs/dock6/parameters/ph4.defn
 ligand_outfile_prefix                                        descriptor.output
 write_footprints                                             yes
 write_hbonds                                                 yes
 write_orientations                                           no
 num_scored_conformers                                        1
 rank_ligands                                                 no

You will now have a file in your directory called descriptor.output_scored.mol2 which can be loaded into Chimera using ViewDock with your original ligand .mol2 file.

=='''II.''de-novo'' growth with PDB 2NNQ (Generic)'''==
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

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.

===Running de novo growth ===
In this section, a de novo growth will be done using the grids generated above and a provided fragment library where the fragments are generated from random drug like ligands.

Go one directory back from multigrid and create a new directory, denovo. Copy the neccesary fragment library files needed into this directory. (scaffold,linker,sidechain,torsion env ..etc) Then create an input file for denovo growth using the lines below. 

 conformer_search_type                                        denovo
 dn_fraglib_scaffold_file                                     fraglib_scaffold.mol2
 dn_fraglib_linker_file                                       fraglib_linker.mol2
 dn_fraglib_sidechain_file                                    fraglib_sidechain.mol2
 dn_user_specified_anchor                                     no
 dn_use_torenv_table                                          yes
 dn_torenv_table                                              fraglib_torenv.dat
 dn_sampling_method                                           graph
 dn_graph_max_picks                                           30
 dn_graph_breadth                                             3
 dn_graph_depth                                               2
 dn_graph_temperature                                         100.0
 dn_pruning_conformer_score_cutoff                            100.0
 dn_pruning_conformer_score_scaling_factor                    1.0
 dn_pruning_clustering_cutoff                                 100.0
 dn_constraint_mol_wt                                         550.0
 dn_constraint_rot_bon                                        15
 dn_constraint_formal_charge                                  2.0
 dn_heur_unmatched_num                                        1
 dn_heur_matched_rmsd                                         2.0
 dn_unique_anchors                                            1
 dn_max_grow_layers                                           4
 dn_max_root_size                                             25
 dn_max_layer_size                                            25
 dn_max_current_aps                                           5
 dn_max_scaffolds_per_layer                                   1
 dn_write_checkpoints                                         yes
 dn_write_prune_dump                                          no
 dn_write_orients                                             no
 dn_write_growth_trees                                        no
 dn_output_prefix                                             output
 use_internal_energy                                          yes
 internal_energy_rep_exp                                      12
 internal_energy_cutoff                                       100.0
 use_database_filter                                          no
 orient_ligand                                                yes
 automated_matching                                           yes
 receptor_site_file                                           ../002.surface_spheres/selected_spheres.sph
 max_orientations                                             1000
 critical_points                                              no
 chemical_matching                                            no
 use_ligand_spheres                                           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                           no
 footprint_similarity_score_secondary                         no
 pharmacophore_score_primary                                  no
 pharmacophore_score_secondary                                no
 descriptor_score_primary                                     yes
 descriptor_score_secondary                                   no
 descriptor_use_grid_score                                    yes
 descriptor_use_multigrid_score                               no
 descriptor_use_pharmacophore_score                           no
 descriptor_use_tanimoto                                      no
 descriptor_use_hungarian                                     no
 descriptor_use_volume_overlap                                yes
 gbsa_zou_score_secondary                                     no
 gbsa_hawkins_score_secondary                                 no
 SASA_score_secondary                                         no
 amber_score_secondary                                        no
 minimize_ligand                                              yes
 minimize_anchor                                              yes
 minimize_flexible_growth                                     yes
 use_advanced_simplex_parameters                              no
 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_anchor_max_iterations                                500
 simplex_grow_max_iterations                                  500
 simplex_grow_tors_premin_iterations                          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

Use the following script (denovo.sh) to submit the job to the cluster.
 #!/bin/bash
 #PBS -l walltime=48:00:00
 #PBS -l nodes=1:ppn=28
 #PBS -q long
 #PBS -N 2nnq_denovo
 #PBS -V
 cd $PBS_O_WORKDIR
 dock6 -i denovo.in -o 2nnq_denovo.out
 
Use the following command to submit the job. 
 qsub denovo.sh