UNIT II – STRUCTURE PREDICTION AND

DRUG DESIGN

Dr. M Indira
Associate Professor
Department of Biotechnology
Vignan University
 SYLLABUS
 Protein structure prediction;
 Introduction to comparative modeling;
 Sequence alignment;
 Constructing and evaluating a comparative model;
 Predicting protein structures by 'threading';
Molecular docking - AUTODOCK/EASYMODELLER
and HEX;
 Structure based de novo ligand design;
 Drug discovery;
 Chemoinformatics; QSAR.
PROTEIN STRUCTURE
PREDICTION
 INTRODUCTION
• Proteins are an important class of
biological macromolecules
which are the polymers of amino
acids.
• Biochemists have distinguished
several levels of structural
organization of proteins. They
are:
– Primary structure
– Secondary structure
– Tertiary structure
– Quaternary structure
 HOMOLOGY MODELLING
INTRODUCTION:
 Homology modeling, also known as comparative modeling of protein
is the technique which allows to construct an unknown atomic-
resolution model of the "target" protein from:
1.Its amino acid sequence and
2.An experimental 3D structure of a related homologous protein (the
"template").

 Prediction of the three dimensional structure of a given protein sequence

i.e. target protein from the amino acid sequence of a homologous
(template) protein for which an X-ray or NMR structure is available based
on an alignment to one or more known protein structures.
 If similarity between the target sequence and the template sequence
is detected, structural similarity can be assumed.
 In general, 30% sequence identity is required to generate an useful
model.
 Homologous proteins
 Portion of 2 proteins with similar amino
acids : Conserved Region
 Highly similar proteins may have same
basic
function
 Homology Modeling
 Comparative modeling of protein
 To predict protein structure based on
known 3D
shape protein as the template
 UNKNOWN STRUCTURE
TEMPLA

?
TE

STRUCTURAL
MODEL
SEQUENCE
ALIGNMENT
 As long as the length of two sequences and the percentage of identical residues fall
in the region marked as “safe” the two sequences are practically guaranteed to adopt
a similar structure.

SEQUENCE SIMILARITY &

STRUCTURAL
SIMILARITY
HOMOLOGY MODELLING
STRUCTURE PREDICTION BY HOMOLOGY
MODELLING
 AN EXAMPLE
 To know the structure of sequence A (150 amino acids long), 1ST of all compare
sequence A to all the sequences of known structures stored in the PDB (using, for
example, BLAST), if a sequence B (300 amino acids long) containing a region of
150 amino acids that match sequence A with 50% identical residues.
 As this match (alignment) clearly falls in the safe zone(50%) , we can simply
take the known structure of sequence B (the template), cut out the fragment
corresponding to the aligned region, mutate those amino acids that differ between
sequences A and B, and finally arrive at our model for structure A. Structure A is
called the target and is of course not known at the time of modeling.

HISTORY
 The first homology modelling studies were done using wire and plastic models of
bonds and atoms as early as the 1960’s. The models were constructed by taking the
coordinates of a known protein structure and modified by hand for those amino
acids that did not match the structure.
 In 1969 David Phillips, Brown and co-workers published the first paper regarding
homology modelling. They modelled -lactalbumin based on the structure of
hen- egg white lysozyme. The sequence identity between these two proteins was
39%.
 STEPS OF HOMOLOGY MODELLING
Protein
1. Template recognition and Sequence
initial alignment
2. Alignment correction Database
Sequence
3. Backbone generation alignment Searches
4. Loop modeling
5. Side chain modeling Secondary
Good
6. Model optimization Structur structure
e prediction
7. Model validation homolog
ue?
Improve
alignment
using
secondary
structure
prediction

Homology
modelling Minimisation

Three
Check dimensional
model structure
 1.Template recognition and initial alignment
 Template recognition & selection involves searching the PDB for
homologous proteins with determined structures.
 The search can be performed using simple sequence alignment programs
such as BLAST or FASTA as the percentage identity between the Target
sequence and a possible template is high enough in the safe zone, to be
detected with these programs.

 To obtain a list of hits-the modeling templates and corresponding

alignments the program compares the query sequence to all the
sequences of known structures in the PDB using mainly two matrices:

1. A residue exchange matrix

2. An alignment matrix .
 2. Alignment correction
 Sometimes it may be difficult to align two sequences in a region where the
percentage sequence identity is very low. One can then use other
sequences from homologous proteins to find a solution.

 For ex: To align the sequence LTLTLTLT with YAYAYAYAY which is nearly
impossible, then only a third sequence, TYTYTYTYT, that aligns easily to
both of them can solve the issue.

 2 is correct, because it leads to a small gap, compared to a huge hole

associated with alignment 1.
 3.BACKBONE GENERATION
 When the alignment is correct, the backbone of the target can be created.
 The coordinates of the template-backbone are copied to the target.
 When the residues are identical, the side-chain coordinates are also
copied.

4.LOOP MODELLING
 After the sequence alignment, there are often regions created by
insertions and deletions that lead to gaps in alignment. These gaps are
modeled by loop modeling, which is less accurate. Currently, two main
techniques are used to approach the problem:
 The database searching method - this involves finding loops from
known protein structures and superimposing them onto the two stem
regions (main chains mostly) of the target protein. Some specialized
programs like FREAD and CODA can be used.
 The ab initio method - this generates many random loops and searches
for one that has reasonably low energy and φ and ψ angles in the
allowable regions in the Ramachandran plot.
 The red loop is modeled with the green
residues as anchor residues. The insertion
of 2 residues results in a longer loop.
 5.Side-Chain Modeling
 This is important in evaluating protein–ligand interactions at active sites
and protein–protein interactions at the contact interface.
 A side chain can be built by searching every possible conformation for
every torsion angle of the side chain to select the one that has the lowest
interaction energy with neighboring atoms.
 A rotamer library can also be used, which has all the favorable side chain
torsion angles extracted from known protein crystal structures.
 6: Model Optimization
 energy minimization procedure on the entire model, by adjusting the
relative position of the atoms so that the overall conformation of the
molecule has the lowest possible energy potential. The goal is to
relieve steric collisions without altering the overall structure.
 Optimization can also be done by Molecular Dynamic Simulation which
moves the atoms toward a global minimum by applying various
stimulation conditions (heating, cooling, considering water molecules)
thus having a better chance at finding the true structure.
 Energy = Stretching Energy +Bending Energy +Torsion Energy +Non-
Bonded Interaction Energy
 7.Model Validation
 Every homology model contains errors. Two main reasons are:
1. The percentage sequence identity between template and target. If it is
greater than 90%, the accuracy of the model can be compared to
crystallographically determined structures & if less than 30% large
error occurs
2. The number of errors in templates
 The final model has to be evaluated for checking the φ–ψ angles,
chirality,
bond lengths, close contacts and also the stereo chemical
properties. Modeling Programs like Modeller, SWISS MODEL,
Schrodinger, 3D- JIGSAW.
 A successful model depends on template selection, algorithm used and the
validation of the model.
 Advantages
 It can find the location of alpha carbons of key residues inside the
folded protein.
 It can help to guide the mutagenesis experiments, or hypothesize
structure-
function relationships.
 The positions of conserved regions of the protein surface can help
identify putative active sites, binding pockets and ligands.

Disadvantages

 Homology models are unable to predict conformations of insertions

or deletions, or side chain positions with a high level of accuracy.
 Homology models are not useful in modeling and ligand docking
studies necessary for the drug designing and development process.
However, it may be helpful for the same, if the sequence identity with
the template is greater than 70%.
http://blast.ncbi.nlm.nih.
gov/
1

3
http://www.ebi.ac.uk/Tools/msa/clusta
lw2/

2
 CPHmodel
 EsyPred3D
 SWISS-
MODEL
http://swissmodel.expasy.
org/
 Swiss-PdbViewer
4.1
 YASARA
 Accelrys DS Viewer
5.0
PDB
YASA
 Verification of
Model
 Verify3D
 ErratPlot
 Ramachandran
Plot
 RAMACHANDRAN PLOT
 In a polypeptide the main chain (N-Calpha)
and (Calpha-C bonds) relatively are free to
rotate. These rotations are represented by the
torsion angles phi (φ) and psi(ψ ), respectively.
 A Ramachandran plot (or a [φ,ψ] plot),
originally developed in 1963 by G. N.
Ramachandran, C. Ramakrishnan, and V.
Sasisekharan,is a way to visualize
backbone dihedral angles ψ against φ of
amino acid residues in protein structure.
A Ramachandran plot can be used:
 One is to show in theory which values, or
conformations, of the ψ and φ angles are
possible for an amino-acid residue in a
protein.
 second is to show the empirical distribution
of datapoints observed in a single structure
in usage for structure validation, or else in a
database of many structures.