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MOE Homlogy Modelling, Building and Evaluating the Protein Structure

Method · September 2018

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SAMEE ULLAH
National Center for Bioinformatics
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Conventions Used in this Tutorial
MOE Window
MOE
Right button bar of the MOE
MOE | RHS
Window
MOE | Footer MOE footer button bar
SE Sequence Editor
SE | Footer Sequence Editor footer bar
DBV Database Viewer

Performing a Homology Search

1. Close the current system using
MOE | File | Close
2. Open the file containing the protein sequence that we will model. The sequence for this
protein is in the MOE sample directory:
MOE | File | Open | $MOE/4muz.fst (your_seq.fst)
3. We will now use the PDB Search application to search for a suitable template for homology
modeling. The search is performed on a database of protein structures and sequences that
have been clustered into families. Open the panel with
MOE | Protein | Search | PDB
Select chain #1 from the Load Chain option list at the top right of the panel to load the
4MUZ.A sequence into the search area and press Search. After starting the search, delete
chain 1 in the Sequence Editor by right-clicking on Chain 1 and selecting Delete — we will be
loading an aligned version of the query after the search.
A two-stage search strategy is used. Firstly, a fast scan is performed to create an initial list of
candidates. Secondly, the list is then narrowed by HMMER, using Hidden Markov Models
(HMM) to determine family membership. The results of the HMMER calculations are shown as
the search progresses. Families are then reported in the Results list as shown.
In this example, the search identifies OMP decarboxylase as a homolog of the target
sequence. Each item in the Results section of the panel represents one or more PDB
structures, possibly augmented by sequence-only entries from the Uniref50 database,
clustered into a family and aligned by a sequence and structure based alignment protocol. The
displayed code represents the PDB structure from within the family with the highest similarity
score to the target sequence.
4. Double-click on PDB_4MUZ.A in the Results area, or select the result and press the
Load Alignment... button. The PDB Search: Load Alignment panel appears, displaying the
pre-aligned family, with the chains sorted by sequence similarity to the query sequence. Note
that the PDB structure PDB_4MUZ.A, which was displayed in the Search results area, is one of
11 PDB structures in this family

5. Select the Load Query Sequence checkbox and press Load All to load these chains into
MOE. In the Sequence Editor, you now see the query sequence (chain 1) as well as all of the
chains from the homologous protein family. Once again, delete chain 2 (4MUZ.A) appearing
under the query before proceeding. You may now close the PDB Search panel.

To annotate the sequences with the secondary structure, on SE | Footer, click

To display the sequences with single letter codes, click

To wrap the sequences to fit into the window, click and scroll down to the
aligned portion of the sequences.
Building a Homology Model
In order to build a homology model, we will align the target sequence to the homologous
structures, decide which chain is to be the template for the model, and then build the model.

1. We will use the Protein Similarity Monitor to compare the sequences. Open the panel with:
SE | Alignment | Similarity
The sequence identity matrix is non-symmetric. The percentage sequence identity is
calculated from the number of identical residues between both chains, divided by the number
of amino acids in the chain corresponding to the cell column.
The matrix shows that the 3THQ.A and 2CZE.A chains are much closer to the query sequence
than the other eight chains.
2. Select chain 4 with the mouse, and then hold the <Shift> key and select chain 11. Then press
the keyboard <Delete> key and click OK to delete these chains.

3. In this case, we are most interested in the residues which differ between the sequences, so we
can highlight residues in the sequence editor which are not conserved between all the chains.

Click SE | Footer | Consensus:

Select the Similarity: * for Conserved Residues check box:

4. Color the residues by the RMS deviation between the atoms using the Residue RMSD button
from the Sequence Editor footer:
SE | Footer | Residues

Tip! Residues with no atoms, like in the sequence-only query chain, are not colored
with the RMSD mode.

Review the aligned sequences by scrolling down in the sequence editor. Note that most of the
residues in the template chains are colored green, showing that they have similar
conformations to each other. Note that no additional superposition is required since the
structures in each PDB family have already been prepared in the database.
Tip! It is good practice to check the positions of the mismatched residues in the
structure. It may be best to choose a template structure with a lower overall
sequence identity, if more of the conserved residues are near a ligand binding site,
or where the residues pack into the middle of the protein. Mismatches can be
accommodated more easily in surface loops, which are more flexible, and have less
steric hindrance.

5. Build the homology model using 3THQ.A as the template structure. Open Homology Model
with
SE | Protein | Homology Model

6. There is only one sequence loaded in MOE with no associated atoms (Chain 1); it is therefore
specified by default as the sequence to be modeled. The Template will default to the first
chain in the system where there are atomic coordinates: Chain 2 (3THQ.A).
Select Open Database Viewer near the top of the Homology Model panel.

7. The currently loaded forcefield is displayed at the bottom of the panel. By default, MOE uses
the Amber10:EHT Forcefield with a Reaction Field (R–Field) treatment of solvation
electrostatics. Verify that these choices are selected, and press Potential Setup to open the
Potential Setup panel to change the settings if needed.
The Fix Hydrogens and Fix Charges buttons in the Potential Setup panel can be ignored.
Homology Model will add or remove hydrogens and lone pairs as necessary, and will assign
partial charges to all atoms.

8. In the Homology Modeling panel press OK. In the output database, promodel.mdb, you can
view the results as they are being calculated.

Notes:
● The entire system is saved prior to modeling as a MOE file specified in the Current System
field. The default filename is promodel.moe.
● The various models will be written to the database specified in the Output Database field.
The default filename is promodel.mdb.
● The Disable C-terminal & N-terminal Outgap Modeling option is, by default, turned on.
When outgaps are ignored, the amino acids of the target sequence that extend beyond (either
before or after) the template sequence in the alignment are clipped off before the model is
built.
● By default, ten independent intermediate models will be built. These different homology
models will be the result of the permutational selection of different loop candidates and
sidechain rotamers.
● The intermediate model which scores best according to the chosen scoring function is chosen
as the final model, subject to optional further energy minimization.
● Once the calculations have finished, the output database will include the ten intermediate
models and a refined final model (entry eleven).
To get a better view of the molecules in the database, you can enlarge the molecular cells in the
Database Viewer. Position the cursor over any one of the cells in the first column (the mol field),
press the left mouse button and drag down and to the right, to enlarge the molecular drawings.
Click the middle mouse button over one of these and drag the mouse to rotate the molecule.
 Comparing the Homology Model with the X-ray
Structure
1. When the calculation finishes, the final, refined model will be loaded in the MOE window.
Note: MOE uses stochastic algorithms in its homology modeling, and results may differ
slightly each time the model is constructed. Therefore, your results and the results shown
here may not be identical.
Open the PDB file with the X-ray structure for 4MUZ:
SE | File | Open | $MOE/sample/mol/4muz.pdb
This will bring up the Load PDB File panel. Keep the default options and press OK.
The 4MUZ structure contains two nearly identical compies of the main chain, as well as other
chains containing ligand and solvent molecules.
2. To compare the homology model with the X-ray structure, we will use
Protein Align/Superpose:
SE | Align/Superpose

Press Align followed by Superpose.

3. In the Sequence Editor, click the - button to the left of the Sequence Editor ruler to hide the
chains with solvent molecules. Open the SE | Footer | Consensus panel and select the
Plot: RMSD check box and deselect the Similarity: * for Conserved Residues check box:

4. Turn on the Selection Synchronize option, so that selecting residues in the Sequence Editor
will select the atoms in those residues in the MOE window, and selecting atoms will select the
residues.
SE | Select | Synchronize
5. Hide the atoms, center the view point and show the protein backbone ribbon using the chain
color:
SE | Footer | Atoms

SE | Footer | Ribbon

to render the final model as shown here.

To simplify the image the second PDB chain

(4MUZ.B) has been hidden in the System Manager. The homology model is shown in
orange.
The X-ray structure is magenta.
Evaluating the Homology Model
It is good practice to examine the geometry of homology models for unusual or unreasonable
features. Serious problems - particularly ones that cluster in one or a few areas of the model - can
suggest a either a problem with the choice or template, or a problem with the alignment of the
target sequence to the template.
1. In the Sequence Editor, select chain 1.
2. Investigate the geometric quality of the model using Protein Geometry from
SE | Protein | Geometry | Phi-Psi Plot
3. From Residues, choose Selected Chains
 The user interface of the Protein Geometry application presents data in the form of plots, such as
the Ramachandran plot shown above, and as tabular lists.
Click on Display: Data and select the outlier(s) in the list, and press Select Atoms at the bottom
of the panel. Now, with the atoms of the outlier residue selected, you may perform operations such
as a fine grain energy minimization in an attempt to obtain more typical phi and psi angles.

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