Template Matching
General remarks
In certain instances it may be useful replace a section of a small
molecule by a new entity. This may be the case when a ligand to a
protein consists of a central template decorated with a set of
substituents that provide binding functionality to the protein.
It may be possible that the substituents are not able to assume an
optimal binding conformation due to geometrical constraints imposed
by the template, or that the template is unsuited for any kind of
reason. For such cases Moloc provides two types of functionality.
Firstly, soft-subset minimization allows the substituents to explore
their optimal binding conformation by treating the template atoms as
a yielding set with reduced force-field parameter values.
Secondly, a set of user-provided templates, a
fragment library, can be searched for
replacement of the original (possibly deformed) template for optimal
substituent attachment positions and exit vectors.
Our eternal (academic) example of dihydrofolate reductase will
provide the protein and methotrexate will be the ligand of which
we consider the central phenyl ring as the template to be replaced.
Soft Subset Minimization
- Read in chain A and methotrexate A as two separate entries from
the file 4dfr.pdb (.../g/p) and set them in the active state (.../a).
- Remove the unnecessary monomer information of methotrexate (.../e/o)
and assure that it is propery protonated (dTp/h)!
- Enter the force field menu (dTp/o, this route avoids setting of
automatic stereo-chemistry conserving constraints in contrast to 'opt').
- Fix the protein (option 's').
- Soften the phenyl ring of methotrexate (option 'Shift s') by
specifying atoms C11 to C16. When leaving the set menu a slider is
presented to specify the reduction factor for force field constants.
Set this slider to its minimum value (0.01).
- Optimize to convergence (option 'o'). The phenyl ring will be badly
distorted.
- Exit from the optimizer, delete the four central atoms of the phenyl
ring (C12, C13, C15, C16) (dTp/d), and save the resulting two-fragment
substituent entry (we call it mtx_sb), e.g. in .mab format (.../s/m).
Preparing a Substituent-Exit-Geometry Entry (Pharmacophore)
The exit geometry of each substituent is given by its point of attachment
and the exit vector of the bond to the first substituent atom. To define
these quantities in Moloc, a pharmacophore entry with directed agons is
well suited.
Manual pharmacophore building (old style):
- Set the protein invisible (.../a) but keep mtx_sb visible.
- Enter the pharmacophore menu (Php) and generate a new entry of
pharmacophore type (option 'n'), which we call mtx. A single agon, LR1,
will appear in the center of the display volume.
- Enter the agon insert menu 'a' (add and delete agons), which is
achieved by picking agon LR1.
- Use option 'b' (add agon directed along a bond) to define two directed
agons by picking the atom pairs C11-C and C14-N10. Set the values of the
first radius and the cone opening angle, to 0.1A and 3 degrees,
respectively. The first radius defines the uncertainty in the attachment
point location, the cone opening angle the uncertainty in the exit vector
direction (the second radius is an alternative but equivalent quantity).
The inserted new agons appear very tiny.
- It may be useful to change the representation of the pharmacophore
(option '.') by setting an atom radius of 1.53 (suboption 'b',
then exit again).
- Delete the initial agon, LR1, (option 'd', pick it with the middle
mouse button) and exit twice to reach the main menu.
- Store the pharmacophore entry to file (.../s/h).
- The pharmacophore builder can also be reached from the template
matching menu (lib/m) with option 'b'.
Automated procedure:
- Keep the (relaxed) ligand (methotrexate) unchanged.
- Go to the template-matching menu (lib/m).
- Select option 'a' and specify the ligand. The 'set' menu appears.
- Specify a user set consisting of the atoms of the template to be
replaced (C11 to C16) and exit from the 'set' menu.
- Now the (two) exit vectors are added consecutively as directed
pharmacophores to a new geometry entry. For each exit vector sliders
are presented to set the ranges in position and direction.
- The pharmacophores receive Moloc extension-types derived from
the atoms on the ligand side.
- Extension types can be changed (specified for old-style
pharmacophores) in option 'e', provided an extension list is available.
Such a list can be specified when reading template libraries (option 'l').
Template Matching
A first (old-style) version of template matching was later improved to
account for selective bond types.
- Old-style templates are Moloc entries in which the points of possible
(or desired) substituent attachment are defined by the heavy atoms carrying
explicit hydrogen atom(s). In the directory moloc/dat a file with a few
very simple templates (tmpl.mab) is provided just for the purpose of
demonstration.
- Moloc templates are structures with terminal atoms carrying
extension-specific labels, single letters that cannot be mistaken for
atoms, followed by a number. For built-in templates unused extensions
assume the atom type they carry in the template, which must not
necessarily be hydrogen.
A default set Ln (n = number) is defined in the file LABEL_moloc. This
file is found in Moloc's 'dat' directory. (By default Moloc looks for files,
the name of which start with 'LABEL', e.g. 'LABEL_my'.)
This file also exemplifies the expected format:
- a list of labels with associated bond orders
- a list of pairs of extensions which can be joined
An example library with some 500 templates up to 10 atoms (DB10.mab)
can be found in Moloc's 'dat' directory. It was derived from DrugBank's
'Small Molecules Structures' file (see link below).
Matching proceeds as follows:
- Enter the template matching menue (lib/m).
- Specify template library (option 'l')
in a new menu. (Old-style just asks for Moloc entries.)
- Specify matching parameters (option 'm'). The parameter f (final
template list) governs the appearence of the results.
- Specify geometry target (option 'g').
- Options 't' and 'v' are optional and concern target pharmacophore
and collision check, respectively.
- With option 's' a substituent molecule (mtx_sb, see above) can be
specified which is required if the f-parameter is set to 2.
This molecules must consist of the substituents up to and including
the attachment atom at the template.
- Option 'd' (do it) starts the search after program has asked for
the template libraries to be searched and for a name (e.g.'m') for the
resulting reoriented (and substituted) templates.
- The remaining options serve to prepare the geometry target.
Examination of Resuts
- Enter Moloc's browse utility (lib/b) and specify the library 'm'
for examination.
- Within option '.' visibility (activity) of all entries not in the
library to be browsed through can be affected. In our case the original
templates can be set to invisible, while the protein should be set in
the active state. This is useful in order to be able to perform energy
evaluations for the identification of possible conflicts of the newly
inserted templates with protein atoms. Furthermore, the pharmacophore
entry, mtx, and, or the substityent entry, mtx_sb, may be set visible
in order to allow to judge the geometrical quality of the substitution.
- The list of proposed substitutions can now be scanned through
forwards (option 'n') or backwards (option 'Shift n').
- Option 'e' allows to examine energies of the currently examined
entry in the context of the protein (when set active).
- Option 'o' generates a duplicate which can then be optimized within
the protein. For this option the protein should be set to fixed the first
time round. Otherwise it will also change its coordinates.