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Epot is the total steric energy which is defined as the difference in energy between
a real molecule and an ideal molecule.
Ebnd, the energy resulting from deforming a bond length from its natural value, is
calculated using Hooke's equation for the deformation of a spring (E = 1/2 Kb(b -
bo)2 where Kb is the force constant for the bond, bo is the equilibrium bond length
and b is the current bond length).
Eang, the energy resulting from deforming a bond angle from its natural value, is
also calculated from Hooke's Law.
Etor is the energy which results from deforming the torsion or dihedral angle.
Folding
3D Structure
First (if structure is known) or second (after structure
prediction) step in a drug design project: find a lead structure
(=small molecule which binds to a given target)
docking problem - predicting the energetically most favorable
complex between a protein and a putative drug molecule
For a given protein structure, one can apply docking
algorithms to virtually search through the space
2 questions:
1. what does the protein-ligand complex look like
2. what is the affinity with respect to other candidates?
Find a set of compounds to start with
- e.g. from inspecting known ligands for a protein (e.g.
substrate in an enzyme)
compounds from a screening experiment of a combinatorial
library (in which there is usually a molecular fragment that is
common between all molecules of the library, the core, and
the fragments attached to the core are R-groups)
compounds from a filtering experiment using other software
from varying other lead structures or known ligands
virtual screening using a fast docking algorithm (typically from
a million molecules)
de novo design using fragments of compounds
=> get several hundred to thousands of ligands to start with
Rigid-body docking algorithms
• Protein and ligand are held fixed in conformational space which reduces
the problem to the search for the relative orientation fo the two molecules
with lowest energy.
Molecular docking
RosettaDOCK
• models physical forces
• Creates a large number of decoys
• degeneracy after clustering is final criterion in selection of
decoys to output
RANDOM START POSITION
Creation of a decoy begins with a random orientation of each
partner and a translation of one partner along the line of
protein centers to create a glancing contact between the
proteins
LOW-RESOLUTION MONTE CARLO SEARCH
Low-resolution representation: N, C, C, O for the backbone
and a “centroid” for the side-chain
One partner is translated and rotated around the surface of the
other through 500 Monte Carlo move attempts
The score terms: A reward for contacting residues, a penalty
for overlapping residues, an alignment score, residue
environment and residue-residue interactions
HIGH-RESOLUTION REFINEMENT
Explicit side-chains are added to the protein backbones using a
rotamer packing algorithm, thus changing the energy surface
An explicit minimization finds the nearest local minimum
accessible via rigid body translation and rotation
Start and Finish positions are compared by the Metropolis
criterion
Before each cycle, the position
of one protein is perturbed by
random translations and by
random rotations
To simultaneously optimize the
side-chain conformations and
the rigid body position, the side-
chain packing and the
minimization operations are
repeated 50 times
COMPUTATIONAL EFFICIENCY
1. The packing algorithm usually varies the conformation of
one residue at a time; rotamer optimization is performed
once every eight cycles
2. Periodically filter to detect and reject inferior decoys
without further refinement
CLUSTERING & PREDICTIONS
Repeat search to create approximately 105 decoys
per target
Cluster best 200 decoys by a hierarchical clustering
algorithm using RMSD
The clusters with the most members become
predictions, ranked by cluster size
Download and install Arguslab in windows
Load a PDB file, practice Arguslab tools
Follow the tutorial at
http://www.arguslab.com/tutorials/tutorial_doc
king_1.htm
Molecular Docking using Argus lab:
Ex : Benzamidine inhibitor docked into Beta Trypsin
Create a binding site from bound ligand
Setting docking
parameters
Analyzing docking results
Polypeptide builder.
The computational molecular docking problem is far
from being solved.
There are two major bottle-necks:
1. The algorithms handle limited flexibility
2. Need selective and efficient scoring functions
Molecular Modeling Applications
Molecular Modeling Applications
Compatibility
Primers used as a pair shall
Primer Pair Matching
work under the same PCR
condition
A melting temperature (Tm) in the range of
~52°C to 65°C
Absence of dimerization capability
Absence of significant hairpin formation (>3
bp)
Lack of secondary priming sites
Low specific binding at the 3' end (ie. lower
GC content to avoid mispriming)
Primer length
GC%
Annealing
3’ complementary between primers
G&C runs at the 3’ end
Palindrome sequences
There shall be one and only one target site in the template
DNA where the primer binds, which means the primer
sequence shall be unique in the template DNA.