2022 Denovo tutorial 2 with PDBID 4ZUD

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De Novo Design

De novo design refers to the process of generating novel ligands in an effort to identify molecules of physiological significance that can be further optimized to become approved drug molecules. The synthesis of thousands of potential drug molecules are done experimentally daily, but with computers, millions of molecules can be computationally modelled and pre-selected for possible synthesis in a fraction of the time it would take to test all possible molecules solely experimentally. With this, scientists are able to direct their attention towards molecules that have the highest probability of imparting a therapeutic effect upon binding to a respective receptor.

This tutorial is the second part of the 2022 DOCK tutorial 2 with PDBID 4ZUD tutorial. You will need the files created in that tutorial to continue with this one!

Make a new directory to organize the files generated in this tutorial:

mkdir 005.denovo

Fragment Library Generation

To create new molecules, we need to begin with the building blocks. For the purposes of speed, we most often use pre-defined molecular fragments that can be arranged/attached in a variety of orientations to create unique structures. Since we have the structure of a ligand that is known to bind the 4ZUD protein, we can generate fragments from that molecule to increase the probability of creating molecules with similar properties to the known ligand.

In an input file:

vim fragment.in

Insert the following:

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.structure/4ZUD_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/dock6.9_release/parameters/vdw_AMBER_parm99.defn
flex_defn_file                                               /gpfs/projects/AMS536/zzz.programs/dock6.9_release/parameters/flex.defn
flex_drive_file                                              /gpfs/projects/AMS536/zzz.programs/dock6.9_release/parameters/flex_drive.tbl
ligand_outfile_prefix                                        fragment.out
write_orientations                                           no
num_scored_conformers                                        1
rank_ligands                                                 no

Run the fragment generation with the following command:

dock6 -i fragment.in -o fragment.out

DOCK should generate six files; three of those files should be mol2's of linker, scaffold, and side chain fragments. You can extract the number of fragments present in each file by running:

grep -wc MOLECULE *.mol2

Fragment Linkers
Fragment Scaffolds
Fragment Side Chains

Focused De Novo Growth

Now we can use the fragments generated in the previous step to allow DOCK to make new molecules that fall within certain rotamers, molecular weights, charges and chemical group compositions. DOCK will generate all possible molecules within the given parameters and that fall within resonable internal energies based on the AMBER van der Waals equation.

In a new directory:

mkdir dn_focused_growth

In an input file:

vim dn_focus.in

Insert the following:

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                                           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                                             dn_focus.out
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                                           yes
grid_score_secondary                                         no

Run this calculation with the following command:

dock6 -i dn_focus.in -o dn_focus.out

A few files will be generated, but the main file with all of the de novo molecules in a mol2 format is dn_focus.out.denovo_build.mol2. If you look at the dn_focus.out file you can see the number of molecules created which should be around 23,000.

Focused De Novo Rescored

Although we've generated a variety of new molecules, all composed of the 4ZUD native ligand fragments, they were created under the energetic constraints of an in silico environment, thus although these new molecules bare similarity to the crystal ligand, many will not be yield viable interaction energies with the 4ZUD receptor.

In order to shorten the list of potential molecules, and to ensure we only focus on ones that form similar interaction with the protein active site relative to the crystal ligand, we can apply a