Difference between revisions of "2024 Denovo tutorial 1 with PDBID 2ITO"

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==Setting Up Your Environment==
 
==Setting Up Your Environment==
−
For this section we will need to create some more directories following this structure:
+
We will add more directories to the ones we used from the traditional virtual screening.
  
−
[[File: updatedStructure.png|center]]
+
  009_deNovoRefinement
 +
  010a_fragLib 
 +
  010b_focusGrowth
 +
  010c_focusReScore
 +
  011_genericDeNovo
  
 
=DeNovo Refinement=
 
=DeNovo Refinement=
−
The DeNovo Refinement algorithm in Dock6.10 is an interesting way to determine the effects on a ligand/protein interaction by changing only part of the small molecule. The part of the ligand we want to experiment with is deleted from the structure, replaced with a dummy atom, and then run through DOCK. The program will try to find which residue can be placed in this now open position that will bind tightly to the protein.
+
DeNovo Refinement can be used to assess the effects upon ligand and protein interactions by chaning a portion of the molecule. We can experiment with an interesting piece of the ligand and delete the remaining portion. This deleted portion needs to be replaced with a dummy atom. Dock will then find which fragments can be placed in this position that will have the least energy.  
  
 
==Setting up the dummy atom==
 
==Setting up the dummy atom==
  
−
For the ligand from #4s0v, we will be removing a terminal ring and looking at what DOCK suggests to replace it with. The steps to do this are: 
+
A dummy atom would be required for setting up the deNovo design process. The dummy atom acts as a starting point for growing the molecule into a new molecule. For our 2TIO system, we would first open the file in Chimera:
−
*Open the ligand minimized mol2 file we generated in the previous tutorial into Chimera.
 
−
*Open the protein into the same session
 
−
*Examine the binding site and choose a residue on the ligand that's pointing towards the inside of the binding site. For our protein this detailed section looks like:
 
  
−
[[File: ligandInSite.png|thumb|center]]
+
[[File: 2tio_ligand_dn.png|thumb|center]]
  
−
We see an imidazole ring pointing towards the binding site so will choose to work with that. Select the protein and hide it from view:
+
In this molecule, we will chose the ring chain to the right to cut it and replace with a dummy atom, since this is the group that falls deeper into the binding site of the protein. The molecule after changing into dummy would mark the dummy atom with a purple atom named Du as shown in Chimera:
  
−
*Place your mouse over the atom connecting the ring to the rest of the ligand and note the atom and number.  In this case it's N4.
+
[[File: 2tiodummy.png|thumb|center]]
−
[[File: atomnumber.png|center]]
 
  
−
*Delete all the atoms from N4 to the end.  Your ligand should now look something like:
+
We were able to change the file to dummy by first deleting the ring group in Chimera by selecting the group and continuously deleting the atoms of the group by Action > Atom > Delete. However we will still leave the atom point where we want to replace it into a dummy point. We will change that by opening the pdb file of the trimmed atom, replacing the name of the atom we want to change to Dummy by editing its name.
−
[[File: DeletedRing.png|center]]
 
  
−
*Save a .mol2 file of your ligand in this configuration.  Make sure to give it a new filename such as 4s0v_denovoRefinement.mol2
+
Shown below here is the text of the pdb file which has already been modified:
−
*Open the .mol2 file.  If you're on a UNIX system, you can use vi; if you're on a PC, you can use textedit.  Locate the atom that will now be changed to a dummy atom:
 
−
[[File: Originalmol2.png|center]]
 
  
−
*Change the atom type to 'Du1' and the bond type to 'Du':
+
[[File: Dummypdb.png|thumb|center]]
−
[[File: Modifiedmol2.png|center]]
 
−
and save the file.
 
  
−
*Open a new session in Chimera and load the modified mol2 file.  The "dummy" atom should now be purple:
+
You can see at atom 23, which was supposed to be a carbon, has been changed the atom name and atom type to "Du". You can use that on your system too but please check in Chimera after editing to see if the atom has been marked purple.
−
[[File: dummyatom.png|thumb|center]]
 
  
−
*scp 4s0v_denovoRefinement.mol2 over to Seawulf and into the 012.denovoRefinement directory. From this point on we will be working on the command line.
+
==Running DeNovo Refinement==
 +
 
 +
After creating the file you would have to upload it onto the server with scp, the file name for the ligand with dummy modified should be named 2tio_denovoRefinement.mol2.
 +
 
 +
Change the working directory the directory for doing the denovo design, there you can create the input file as followed:
 +
 
 +
  vi denovoRefinement.in
 +
 
 +
There you can create the parameter for the input file as followed:
  
−
==Running DeNovo Refinement==
+
  conformer_search_type                                        denovo
 +
  dn_fraglib_scaffold_file                                    /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_scaffold.mol2
 +
  dn_fraglib_linker_file                                      /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_linker.mol2
 +
  dn_fraglib_sidechain_file                                    /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_sidechain.mol2
 +
  dn_user_specified_anchor                                    yes
 +
  dn_fraglib_anchor_file                                      2tio_ligand_denovo.mol2
 +
  dn_torenv_table                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_torenv.dat
 +
  dn_name_identifier                                          2tio_denovo
 +
  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                    2.0
 +
  dn_pruning_clustering_cutoff                                100.0
 +
  dn_mol_wt_cutoff_type                                        soft
 +
  dn_upper_constraint_mol_wt                                  1000.0
 +
  dn_lower_constraint_mol_wt                                  0.0
 +
  dn_mol_wt_std_dev                                            35.0
 +
  dn_constraint_rot_bon                                        15
 +
  dn_constraint_formal_charge                                  5
 +
  dn_heur_unmatched_num                                        1
 +
  dn_heur_matched_rmsd                                        2.0
 +
  dn_unique_anchors                                            1
 +
  dn_max_grow_layers                                          1
 +
  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                                            2tio_denovo_output
 +
  use_internal_energy                                          yes
 +
  internal_energy_rep_exp                                      12
 +
  internal_energy_cutoff                                      100.0
 +
  use_database_filter                                          no
 +
  orient_ligand                                                no
 +
  bump_filter                                                  no
 +
  score_molecules                                              yes
 +
  contact_score_primary                                        no
 +
  grid_score_primary                                          yes
 +
  grid_score_rep_rad_scale                                    1
 +
  grid_score_vdw_scale                                        1
 +
  grid_score_es_scale                                          1
 +
  grid_score_grid_prefix                                      ../003.gridbox/grid
 +
  minimize_ligand                                              yes
 +
  minimize_anchor                                              no
 +
  minimize_flexible_growth                                    yes
 +
  use_advanced_simplex_parameters                              no
 +
  simplex_max_cycles                                          1
 +
  simplex_score_converge                                      0.1
 +
  simplex_cycle_converge                                      1
 +
  simplex_trans_step                                          1
 +
  simplex_rot_step                                            0.1
 +
  simplex_tors_step                                            10
 +
  simplex_grow_max_iterations                                  250
 +
  simplex_grow_tors_premin_iterations                          0
 +
  simplex_random_seed                                          0
 +
  simplex_restraint_min                                        yes
 +
  simplex_coefficient_restraint                                10
 +
  atom_model                                                  all
 +
  vdw_defn_file                                                /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/vdw_de_novo.defn
 +
  flex_defn_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
 +
  flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl
 +
 
 +
After finishing the parameter files, you can start running denovo design:
 +
 
 +
  dock6 -i denovoRefinement.in -o denovoRefinement.out
 +
 
 +
Once the job has completed you will see the following new files in your directory:
  
 +
*2tio_denovo_output.anchor_1.root_layer_1.mol2
 +
*2tio_denovo_output.denovo_build.mol2
 +
*denovoRefinement.out
  
 
==Viewing New Molecules==
 
==Viewing New Molecules==
 +
 +
In order to view the denovo docking result, you can download the 2tio_denovo_output.denovo_build.mol2 to your local computer and view that file in Chimera with ViewDock in Chimera (under Tools > Surface/Binding Analysis > ViewDock). Following is an example output:
 +
 +
[[File: 2tio_dn_1.png|center|600px]]
 +
 +
[[File: 2tio_dn_2.png|center|600px]]
  
 
=Focused DeNovo Design=
 
=Focused DeNovo Design=
 +
 +
The subsequent two types of de novo design share a resemblance in which, with this approach, a specific structure is chosen to "anchor" the ligand to the protein. DOCK will then explore this structure throughout the protein to identify the optimal binding site. From this anchor point, a new ligand is developed. The distinction between focused and generic de novo design lies in the specific residues DOCK employs to build this new ligand.
  
 
==Fragment Library Generation==
 
==Fragment Library Generation==
 +
 +
To begin focus deNovo design, the first step is to construct a fragment library that is compatible with the ligand-complex system that one is interested in. In other to do so, we will use the original ligand along with the selected sphere from the surface step as a template for generating the fragment library and the algorithm to do so is embedded in DOCK. Thus, we will be gin by changing the working directory to the 013a.fragLib.in.
 +
 +
Here we will initiate an input file for the program:
 +
 +
  vi fragLib.in
 +
 +
The following should be the input parameters for the file:
 +
 +
  conformer_search_type                                        flex
 +
  write_fragment_libraries                                    yes
 +
  fragment_library_prefix                                      2tio_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.structure/2tio_ligand_hydrogens.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/dock6.10/parameters/vdw_de_novo.defn
 +
  flex_defn_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
 +
  flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl
 +
  ligand_outfile_prefix                                        2tio_focused
 +
  write_orientations                                          no
 +
  num_scored_conformers                                        1
 +
  write_conformations                                          no
 +
  cluster_conformations                                        yes
 +
  cluster_rmsd_threshold                                      2.0
 +
  rank_ligands                                                no
 +
 +
After finish compiling the input file for fragment generation, we can start running the program:
 +
 +
  dock6 -i fragLib.in -o fragLib.out
 +
 +
The output file should be contain the following, note that if we fail to generate sufficient fragment library, there would not be enough input file for sequel steps:
 +
 +
*2tio_focused_scored.mol2
 +
*2tio_fragLib_linker.mol2
 +
*2tio_fragLib_rigid.mol2
 +
*2tio_fragLib_scaffold.mol2
 +
*2tio_fragLib_sidechain.mol2
 +
*2tio_fragLib_torenv.dat
  
 
==DeNovo Design==
 
==DeNovo Design==
 +
 +
In this next step we will be navigating to the working directory for deNovo design using the Focus algorithm. The name of the directory for working would be 013b.foucusGrowth. As a sequel step from identifying the fragment library in the previous step, it would be required that you already achieved the library fragments set and store it correctly in the 013a.fragLib.
 +
 +
Initiating the input file with vi:
 +
 +
vi deNovoFocus.in
 +
 +
Insert the following content for the input:
 +
 +
conformer_search_type                                        denovo
 +
dn_fraglib_scaffold_file                                    ../013a.fragLib/2tio_fragLib_scaffold.mol2
 +
dn_fraglib_linker_file                                      ../013a.fragLib/2tio_fragLib_linker.mol2
 +
dn_fraglib_sidechain_file                                    ../013a.fragLib/2tio_fragLib_sidechain.mol2
 +
dn_user_specified_anchor                                    no
 +
dn_use_torenv_table                                          yes
 +
dn_torenv_table                                              ../013a.fragLib/2tio_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                                            2
 +
dn_max_grow_layers                                          9
 +
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                                        yes
 +
dn_output_prefix                                            2tio_focused
 +
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/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                                          yes
 +
grid_score_secondary                                        no
 +
grid_score_rep_rad_scale                                    1
 +
grid_score_vdw_scale                                        1
 +
grid_score_es_scale                                          1
 +
grid_score_grid_prefix                                      ../003.gridbox/grid
 +
multigrid_score_secondary                                    no
 +
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
 +
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/dock6.10/parameters/vdw_de_novo.defn
 +
flex_defn_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
 +
flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl
 +
 +
Start running the deNovo focus algorithm with the input as:
 +
 +
dock6 -i deNovoFocus.in -o deNovoFocus.out
 +
 +
After running the command, you should leave your program run until it executes. The program could not be run on server with mpi run. After you have successfully run the command, please check the directory output file to see if there have been any errors and the directory should contain multiple mol2 files in which the build.mol2 would be summarizing the molecules that have been built with denovo design algorithm.
  
 
==ReScoring Designed Molecules==
 
==ReScoring Designed Molecules==
−
   
+
 
 +
In other to make the multiples molecules suggested in the last step become more informative on which molecule would be the most likely to become the drug lead, in this step, the molecule set would be re-scored so that each would be compared with the binding of the original binding ligand to see if they would govern a similar effect to the original binder. The algorithm used for this scoring would be footprint similarity score, to begin, we would navigate to the directory designated for this section 013c.focusReScore. Here please initiate by creating the input file:
 +
 
 +
  vi focusReScore.in
 +
 
 +
Insert the following content for the input:
 +
 
 +
conformer_search_type                                        rigid
 +
use_internal_energy                                          yes
 +
internal_energy_rep_exp                                      12
 +
internal_energy_cutoff                                      100.0
 +
ligand_atom_file                                            ../013b.focusGrowth/2tio_focused.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      ../004.energy_min/2tio.lig.min_scored.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.structure/2tio_protein_hydrogens_charges.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                      ../004.energy_min/2tio.lig.min_scored.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                          ../004.energy_min/2tio.lig.min_scored.mol2
 +
descriptor_hms_score_ref_filename                            ../004.energy_min/2tio.lig.min_scored.mol2
 +
descriptor_hms_score_matching_coeff                          -5
 +
descriptor_hms_score_rmsd_coeff                              1
 +
descriptor_volume_score_reference_mol2_filename              ../004.energy_min/2tio.lig.min_scored.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/dock6.10/parameters/vdw_de_novo.defn
 +
flex_defn_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
 +
flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl
 +
chem_defn_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/chem.defn
 +
pharmacophore_defn_file                                      /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/ph4.defn
 +
ligand_outfile_prefix                                        2tio_focus_rescore
 +
write_footprints                                            yes
 +
write_hbonds                                                yes
 +
write_orientations                                          no
 +
num_scored_conformers                                        1
 +
rank_ligands                                                no
 +
 
 +
Start running the rescore algorithm with the input as:
 +
 
 +
dock6 -i focusReScore.in -o focusReScore.out
 +
 
 +
If you have successfully computed the simulation you should receive the output results as followed:
 +
 4s0v_focus_rescore_footprint_scored.txt
 +
 4s0v_focus_rescore_hbond_scored.txt
 +
 4s0v_focus_rescore_scored.mol2
 +
 
 +
These are the sets of molecules suggested by deNovo being ranked based on the footprint scores (as mentioned above), and specifically with the hydrogen bonding scores. In other to view the results, you would need to load the result mol2 attached down to your local computer and view that with Chimera.
 +
 
 +
Below is loading the mol2 with ViewDock and ranking the file based on the footprint similarity score:
 +
 
 +
[[File:Focus1png.png]]
 +
 
 +
[[File:Focus2png.png]]
 +
 
 
=Generic DeNovo Design=
 
=Generic DeNovo Design=
 +
 +
For generic denovo design, we are not limiting DOCK to build new molecules using fragments present in the original ligand. Instead, we will be taking fragments from a fragment library.
 +
 +
Create the input file:
 +
 +
  vi 2ito_generic.in
 +
 +
The file should contain these lines:
  
 
   conformer_search_type                                        denovo
 
   conformer_search_type                                        denovo
Line 134: Line 495:
 
   flex_defn_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
 
   flex_defn_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
 
   flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl
 
   flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl
 +
 +
Create a slurm file to run this:
 +
 +
  #!/bin/bash
 +
  #SBATCH --job-name=2ito_generic
 +
  #SBATCH --ntasks=28
 +
  #SBATCH --nodes=1
 +
  #SBATCH --time=48:00:00
 +
  #SBATCH -p long-28core
 +
  dock6 -i 2ito_generic.in
 +
 +
After the program has run, you can use the following command to see how many ligands has been generated. The program has made 368 potential ligands.
 +
 +
  grep MOLECULE 2ito_generic.denovo_build.mol2 | wc -l
 +
 +
 +
The output file we are going to move to our local computer is 2ito_generic.denovo_build.mol2. We can see the ligands the program has created.
 +
 +
Go into Chimera and open the molecule on ViewDock. (Tools >> Surface/Binding Analysis >> ViewDock)
 +
 +
Use Column >> Show >> Grid_Score to find the ligand with the lowest energy. The ligand with the lowest score has a score of -93.318.
 +
 +
[[File: 2ITO_genericDenovo_lowest_gridscore.png|thumb|center]]
 +
[[File:2ITO_genericDeNovo_ligands_gridScores.png|thumb|center]]
 +
 +
 +
We can compare this with the energy minimized ligand and see how it is positioned differently compared to the generated ligand (denovo_designed ligand is brown, energy minimized ligand is blue). This original energy minimized ligand has a grid score of -71.423.
 +
 +
[[File: LowestScoreLigand original.png|thumb|center]]
 +
 +
 +
We can also observe how the designed ligand and the original ligand interact with the receptor.
 +
 +
[[File: Lowest original 2ITOprotein.png|thumb|center]]

Latest revision as of 09:43, 6 May 2024

Introduction

DeNovo Design aims to create a new ligand from scratch. It is incrementally designed bad added known structures of bioacitve compounds. The end result will hopefully produce a new ligand that can bind more tightly to a protein receptor than the current ligand can. There are three methods that will be introduced in this tutorial:

  1. Generic DeNovo Design
  2. Focused Fragment Design
  3. DeNovo Refinement

We will do denovo design with PDB: 2ITO.


Setting Up Your Environment

We will add more directories to the ones we used from the traditional virtual screening. 

 009_deNovoRefinement
 010a_fragLib   
 010b_focusGrowth
 010c_focusReScore
 011_genericDeNovo

DeNovo Refinement

DeNovo Refinement can be used to assess the effects upon ligand and protein interactions by chaning a portion of the molecule. We can experiment with an interesting piece of the ligand and delete the remaining portion. This deleted portion needs to be replaced with a dummy atom. Dock will then find which fragments can be placed in this position that will have the least energy.

Setting up the dummy atom

A dummy atom would be required for setting up the deNovo design process. The dummy atom acts as a starting point for growing the molecule into a new molecule. For our 2TIO system, we would first open the file in Chimera:

2tio ligand dn.png

In this molecule, we will chose the ring chain to the right to cut it and replace with a dummy atom, since this is the group that falls deeper into the binding site of the protein. The molecule after changing into dummy would mark the dummy atom with a purple atom named Du as shown in Chimera:

2tiodummy.png

We were able to change the file to dummy by first deleting the ring group in Chimera by selecting the group and continuously deleting the atoms of the group by Action > Atom > Delete. However we will still leave the atom point where we want to replace it into a dummy point. We will change that by opening the pdb file of the trimmed atom, replacing the name of the atom we want to change to Dummy by editing its name.

Shown below here is the text of the pdb file which has already been modified:

Dummypdb.png

You can see at atom 23, which was supposed to be a carbon, has been changed the atom name and atom type to "Du". You can use that on your system too but please check in Chimera after editing to see if the atom has been marked purple.

Running DeNovo Refinement

After creating the file you would have to upload it onto the server with scp, the file name for the ligand with dummy modified should be named 2tio_denovoRefinement.mol2.

Change the working directory the directory for doing the denovo design, there you can create the input file as followed: 

 vi denovoRefinement.in

There you can create the parameter for the input file as followed: 

 conformer_search_type                                        denovo
 dn_fraglib_scaffold_file                                     /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_scaffold.mol2
 dn_fraglib_linker_file                                       /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_linker.mol2
 dn_fraglib_sidechain_file                                    /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_sidechain.mol2
 dn_user_specified_anchor                                     yes
 dn_fraglib_anchor_file                                       2tio_ligand_denovo.mol2
 dn_torenv_table                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_torenv.dat
 dn_name_identifier                                           2tio_denovo
 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                    2.0
 dn_pruning_clustering_cutoff                                 100.0
 dn_mol_wt_cutoff_type                                        soft
 dn_upper_constraint_mol_wt                                   1000.0
 dn_lower_constraint_mol_wt                                   0.0
 dn_mol_wt_std_dev                                            35.0
 dn_constraint_rot_bon                                        15
 dn_constraint_formal_charge                                  5
 dn_heur_unmatched_num                                        1
 dn_heur_matched_rmsd                                         2.0
 dn_unique_anchors                                            1
 dn_max_grow_layers                                           1
 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                                             2tio_denovo_output
 use_internal_energy                                          yes
 internal_energy_rep_exp                                      12
 internal_energy_cutoff                                       100.0
 use_database_filter                                          no
 orient_ligand                                                no
 bump_filter                                                  no
 score_molecules                                              yes
 contact_score_primary                                        no
 grid_score_primary                                           yes
 grid_score_rep_rad_scale                                     1
 grid_score_vdw_scale                                         1
 grid_score_es_scale                                          1
 grid_score_grid_prefix                                       ../003.gridbox/grid
 minimize_ligand                                              yes
 minimize_anchor                                              no
 minimize_flexible_growth                                     yes
 use_advanced_simplex_parameters                              no
 simplex_max_cycles                                           1
 simplex_score_converge                                       0.1
 simplex_cycle_converge                                       1
 simplex_trans_step                                           1
 simplex_rot_step                                             0.1
 simplex_tors_step                                            10
 simplex_grow_max_iterations                                  250
 simplex_grow_tors_premin_iterations                          0
 simplex_random_seed                                          0
 simplex_restraint_min                                        yes
 simplex_coefficient_restraint                                10
 atom_model                                                   all
 vdw_defn_file                                                /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/vdw_de_novo.defn
 flex_defn_file                                               /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
 flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl

After finishing the parameter files, you can start running denovo design: 

 dock6 -i denovoRefinement.in -o denovoRefinement.out

Once the job has completed you will see the following new files in your directory:

  • 2tio_denovo_output.anchor_1.root_layer_1.mol2
  • 2tio_denovo_output.denovo_build.mol2
  • denovoRefinement.out

Viewing New Molecules

In order to view the denovo docking result, you can download the 2tio_denovo_output.denovo_build.mol2 to your local computer and view that file in Chimera with ViewDock in Chimera (under Tools > Surface/Binding Analysis > ViewDock). Following is an example output:

2tio dn 1.png
2tio dn 2.png

Focused DeNovo Design

The subsequent two types of de novo design share a resemblance in which, with this approach, a specific structure is chosen to "anchor" the ligand to the protein. DOCK will then explore this structure throughout the protein to identify the optimal binding site. From this anchor point, a new ligand is developed. The distinction between focused and generic de novo design lies in the specific residues DOCK employs to build this new ligand.

Fragment Library Generation

To begin focus deNovo design, the first step is to construct a fragment library that is compatible with the ligand-complex system that one is interested in. In other to do so, we will use the original ligand along with the selected sphere from the surface step as a template for generating the fragment library and the algorithm to do so is embedded in DOCK. Thus, we will be gin by changing the working directory to the 013a.fragLib.in.

Here we will initiate an input file for the program: 

 vi fragLib.in

The following should be the input parameters for the file: 

 conformer_search_type                                        flex
 write_fragment_libraries                                     yes
 fragment_library_prefix                                      2tio_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.structure/2tio_ligand_hydrogens.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/dock6.10/parameters/vdw_de_novo.defn
 flex_defn_file                                               /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
 flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl
 ligand_outfile_prefix                                        2tio_focused
 write_orientations                                           no
 num_scored_conformers                                        1
 write_conformations                                          no
 cluster_conformations                                        yes
 cluster_rmsd_threshold                                       2.0
 rank_ligands                                                 no

After finish compiling the input file for fragment generation, we can start running the program: 

 dock6 -i fragLib.in -o fragLib.out

The output file should be contain the following, note that if we fail to generate sufficient fragment library, there would not be enough input file for sequel steps:

  • 2tio_focused_scored.mol2
  • 2tio_fragLib_linker.mol2
  • 2tio_fragLib_rigid.mol2
  • 2tio_fragLib_scaffold.mol2
  • 2tio_fragLib_sidechain.mol2
  • 2tio_fragLib_torenv.dat

DeNovo Design

In this next step we will be navigating to the working directory for deNovo design using the Focus algorithm. The name of the directory for working would be 013b.foucusGrowth. As a sequel step from identifying the fragment library in the previous step, it would be required that you already achieved the library fragments set and store it correctly in the 013a.fragLib.

Initiating the input file with vi: 

vi deNovoFocus.in

Insert the following content for the input: 

conformer_search_type                                        denovo
dn_fraglib_scaffold_file                                     ../013a.fragLib/2tio_fragLib_scaffold.mol2
dn_fraglib_linker_file                                       ../013a.fragLib/2tio_fragLib_linker.mol2
dn_fraglib_sidechain_file                                    ../013a.fragLib/2tio_fragLib_sidechain.mol2
dn_user_specified_anchor                                     no
dn_use_torenv_table                                          yes
dn_torenv_table                                              ../013a.fragLib/2tio_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                                            2
dn_max_grow_layers                                           9
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                                        yes
dn_output_prefix                                             2tio_focused
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/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                                           yes
grid_score_secondary                                         no
grid_score_rep_rad_scale                                     1
grid_score_vdw_scale                                         1
grid_score_es_scale                                          1
grid_score_grid_prefix                                       ../003.gridbox/grid
multigrid_score_secondary                                    no
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
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/dock6.10/parameters/vdw_de_novo.defn
flex_defn_file                                               /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl

Start running the deNovo focus algorithm with the input as: 

dock6 -i deNovoFocus.in -o deNovoFocus.out

After running the command, you should leave your program run until it executes. The program could not be run on server with mpi run. After you have successfully run the command, please check the directory output file to see if there have been any errors and the directory should contain multiple mol2 files in which the build.mol2 would be summarizing the molecules that have been built with denovo design algorithm.

ReScoring Designed Molecules

In other to make the multiples molecules suggested in the last step become more informative on which molecule would be the most likely to become the drug lead, in this step, the molecule set would be re-scored so that each would be compared with the binding of the original binding ligand to see if they would govern a similar effect to the original binder. The algorithm used for this scoring would be footprint similarity score, to begin, we would navigate to the directory designated for this section 013c.focusReScore. Here please initiate by creating the input file: 

vi focusReScore.in

Insert the following content for the input:

conformer_search_type rigid 

use_internal_energy                                          yes
internal_energy_rep_exp                                      12
internal_energy_cutoff                                       100.0
ligand_atom_file                                             ../013b.focusGrowth/2tio_focused.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       ../004.energy_min/2tio.lig.min_scored.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.structure/2tio_protein_hydrogens_charges.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                       ../004.energy_min/2tio.lig.min_scored.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                          ../004.energy_min/2tio.lig.min_scored.mol2
descriptor_hms_score_ref_filename                            ../004.energy_min/2tio.lig.min_scored.mol2
descriptor_hms_score_matching_coeff                          -5
descriptor_hms_score_rmsd_coeff                              1
descriptor_volume_score_reference_mol2_filename              ../004.energy_min/2tio.lig.min_scored.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/dock6.10/parameters/vdw_de_novo.defn
flex_defn_file                                               /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl
chem_defn_file                                               /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/chem.defn
pharmacophore_defn_file                                      /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/ph4.defn

ligand_outfile_prefix 2tio_focus_rescore 

write_footprints                                             yes
write_hbonds                                                 yes
write_orientations                                           no
num_scored_conformers                                        1
rank_ligands                                                 no

Start running the rescore algorithm with the input as: 

dock6 -i focusReScore.in -o focusReScore.out

If you have successfully computed the simulation you should receive the output results as followed:  4s0v_focus_rescore_footprint_scored.txt  4s0v_focus_rescore_hbond_scored.txt  4s0v_focus_rescore_scored.mol2

These are the sets of molecules suggested by deNovo being ranked based on the footprint scores (as mentioned above), and specifically with the hydrogen bonding scores. In other to view the results, you would need to load the result mol2 attached down to your local computer and view that with Chimera.

Below is loading the mol2 with ViewDock and ranking the file based on the footprint similarity score:

Focus1png.png

Focus2png.png

Generic DeNovo Design

For generic denovo design, we are not limiting DOCK to build new molecules using fragments present in the original ligand. Instead, we will be taking fragments from a fragment library.

Create the input file: 

 vi 2ito_generic.in

The file should contain these lines: 

 conformer_search_type                                        denovo
 dn_fraglib_scaffold_file                                     /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_scaffold.mol2
 dn_fraglib_linker_file                                       /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_linker.mol2
 dn_fraglib_sidechain_file                                    /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_sidechain.mol2
 dn_user_specified_anchor                                     no
 dn_torenv_table                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/fraglib_torenv.dat
 dn_name_identifier                                           2ito_generic
 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                    2.0
 dn_pruning_clustering_cutoff                                 100.0
 dn_mol_wt_cutoff_type                                        soft
 dn_upper_constraint_mol_wt                                   550.0
 dn_lower_constraint_mol_wt                                   0.0
 dn_mol_wt_std_dev                                            35.0
 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                                           9
 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                                             2ito_generic
 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                                           /gpfs/projects/AMS536/2024/students/group_1_2ITO/002_surface_spheres/2ITO.sph
 max_orientations                                             1000
 critical_points                                              no
 chemical_matching                                            no
 use_ligand_spheres                                           no
 bump_filter                                                  no
 score_molecules                                              yes
 contact_score_primary                                        no
 grid_score_primary                                           yes
 grid_score_rep_rad_scale                                     1
 grid_score_vdw_scale                                          1
 grid_score_es_scale                                           1
 grid_score_grid_prefix                                       /gpfs/projects/AMS536/2024/students/group_1_2ITO/003_gridbox/grid
 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                                  250
 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/dock6.10/parameters/vdw_de_novo.defn
 flex_defn_file                                               /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex.defn
 flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.10/parameters/flex_drive.tbl

Create a slurm file to run this: 

 #!/bin/bash
 #SBATCH --job-name=2ito_generic
 #SBATCH --ntasks=28
 #SBATCH --nodes=1
 #SBATCH --time=48:00:00
 #SBATCH -p long-28core
 dock6 -i 2ito_generic.in

After the program has run, you can use the following command to see how many ligands has been generated. The program has made 368 potential ligands. 

 grep MOLECULE 2ito_generic.denovo_build.mol2 | wc -l


The output file we are going to move to our local computer is 2ito_generic.denovo_build.mol2. We can see the ligands the program has created.

Go into Chimera and open the molecule on ViewDock. (Tools >> Surface/Binding Analysis >> ViewDock)

Use Column >> Show >> Grid_Score to find the ligand with the lowest energy. The ligand with the lowest score has a score of -93.318.

2ITO genericDenovo lowest gridscore.png
2ITO genericDeNovo ligands gridScores.png


We can compare this with the energy minimized ligand and see how it is positioned differently compared to the generated ligand (denovo_designed ligand is brown, energy minimized ligand is blue). This original energy minimized ligand has a grid score of -71.423.

LowestScoreLigand original.png


We can also observe how the designed ligand and the original ligand interact with the receptor.

Lowest original 2ITOprotein.png
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