Farhan Ahmed
Protein Modeling Project
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Hetero Protein Complex: Manganese Protoporphyrin IX-reconstituted Ovine
Prostaglandin H2 Synthase-1 Complexed with Flurbiprofen
Protein: PDB ID 2ayl
Hetero-compound: Flurbiprofen,
C15H13FO2, Het code FLP.
IUPAC Name: 2-(3-fluoro-4-phenyl-phenyl)
propanoic acid
The RCSB PDB database was
accessed in order to study the heteroImage 1: After acquiring the PDB file from
RCSB PDB. The hetero protein complex structure
as displayed in DS Visualizer.
compound, flurbiprofen. Through the
database the three letter code was found to be
flp, while the protein complex was given the code 2ayl.
The PDB file was
downloaded and saved on to the
computer. The PDB file was
opened by using the program
DS Visualizer. The structure of
the protein complex was on
display once the file was
opened, as seen in image 1.
Image 2: Results of the protein
extraction. This image does not show the
right hybridization or the hydrogen atoms
bonded to the compound.
The hetero compound was located in the
interior of the protein. In order to extract the hetero
compound, the extra groups were removed. The first
groups to be removed were those of water, followed by the protein chains not complexed to the
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Protein Modeling Project
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active compound. The hetero compound was transferred to a different window by copying and
pasting it to a new window. The results of the protein extraction are shown in image 2. This
image does not contain the proper
hybridization or the correct number
of Hydrogen atoms. The instructions
from the class website were followed
in order to make the proper
adjustments. The results of the
change are displayed in image 3.
The program Chem. Sketch
Image 3: The corrected hybridization of the hetero-compound
and the addition of Hydrogen atoms to the hetero compound.
was used in order to draw the Lewis
structure of the hetero compound, the results
of which are on display in image 4. In DSV, the protein-hetero compound complex was
displayed, with the protein shown in solid ribbon style and the bound hetero compound in CPK
style. These results are shown in image 6.
CH3
O
O
F
Image 4: The Lewis structure of Flurbiprofen
was drawn using the program Chem. Sketch.
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Protein Modeling Project
Image 6: the protein-hetero compound complex was displayed,
with the protein shown in solid ribbon style and the bound hetero
compound in CPK style.
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Farhan Ahmed
Protein Modeling Project
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Part II: Steric energy calculations, superimposing the extracted and energy-minimized hetero
compounds.
Image 5: Before MM2 minimization.
Image 6: After MM2 minimization
This value is the steric energy of the compound when it is complexed within the protein. It is
probably not a local or global energy minimum as found for the same compound in the gas phase
where there are no intermolecular interactions.
Energy terms
Stretch
Bend
Stretch-Bend
Torsion
Non-1,4 VDW
1,4 VDW
Charge/Charge
Charge/Dipole
Dipole/Dipole
Total Energy
Complex of
Prostaglandin H2
synthase-1 with
Flurbiprofen
Energy minimization Difference
36.9628
0.4151
-36.5477
2.52
3.2174
0.6974
-0.0555
-0.1519
-0.0964
-11.1878
-30.5571
-19.3693
-1.3863
-2.6047
-1.2184
7.8891
4.1394
-3.7497
0
0
0
-1.5053
-1.6425
-0.1372
0.7073
0.6798
-0.0275
33.9444
-26.5046
-60.449
Table 1: The Energy components and total energy for flurbiprofen
in the complex form and the minimized form. Column three shows
the energy difference between each term.
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Protein Modeling Project
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Image 5 shows the compound as it appears in the hetero-protein complex, while image 6
shows the conformation of flurbiprofen in its most stable gas phase. The most stable gas phase
was achieved after performing an energy minimization in the program Chem. 3D. Table 1 was
also calculated using Chem. 3D, it shows the energy values of each of the energy component
terms as well as the total energy for each of conformation. The difference in the energies of the
two conformations is represented in the third column of table 1. A total energy difference of 60.449kcal/mol is computed between the two conformations. The difference in energies is a
result of steric clashes taking place in Image 5 of flurbiprofen.
After obtaining the energy values of each of the energy component it was determined
that the total energy difference between the two conformations was 60.449kcal/mol. It can
then be said that the complexed flurbiprofen is in a high energy conformation as compared to
the gas phase. Further examination of the data shows that the “stretch” and the “torsion”
values display the greatest difference in the conformational analysis data; therefore the main
contribution of energy difference is a result of these two values. The other terms produce
minor differences in energy values and do not contribute much to the difference in total
energy. The Charge/Charge term produces a value of zero in both conformations. This is due to
the fact that this term describes electrostatic interactions between charged atoms and functional
groups in the molecule and the only charged group in the molecule is the carboxylate group.
An examination of image 5 and image 6 reveals the conformational differences that result
in the energy difference. In image 5 the two rings are in a twisted formation, which could disturb
the overlapping of the pie-orbital resulting in the inability to share electrons effectively. As table
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Protein Modeling Project
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1 indicates the “torsion” is one value that contributes significantly to the difference in energy.
The twisting of the rings could
result in increased torsional strain
which leads to a high energy
conformation.
The difference in structure of
the flurbiprofen is easier to see
when the two structures are placed
on top of each other. Although there
are many points of similarities in the
Image 7: The structures of flurbiprofen at different energy
conformations are placed on top of each other. One
structure is in the plain while the other is not, this adds to
the torsion strain.
structures, it is clear that there is a difference in the rings of the structures. In one of the
structures the ring system is in the plain of the paper, while in the other it is facing outward. This
difference in structure contributes to the energy term “torsion.”
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Farhan Ahmed
Protein Modeling Project
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Part III: Protein-Ligand
Interactions
In Part three of this
assignment, a protein wiring
diagram was accessed from
the PDBSum database. The
wiring diagram obtained can
be seen in image 8. The
wiring diagram displays the
numbered amino acid
sequence, of the protein using
one-letter codes. It also
includes information about
the secondary structure,
including alpha helices, beta
sheets, hairpin turns, and
disulfide bonds. The red dots
above certain amino acid
residues indicate interactions
of the protein with a ligand,
while the blue dots represent
Image 8: The protein wiring diagram
obtained from the PDBSum database.
an interaction with a metal. A
red box indicates a residue with catalytic properties.
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Farhan Ahmed
Protein Modeling Project
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In order to determine which residues interact with the complexed flurbiprofen molecule,
a Lig-Plot was obtained from the same the PDBSum website. A Lig-Plot is a two-dimensional
chart that represents the ligand interaction. The Lig-Plot obtained for flurbiprofen is shown in
image 9. A Lig-Plot shows the amino acid residues involved in complexing the heterocompound. Since the Lig-Plot is a two dimensional chart, it is difficult to know the placement of
the amino acids. A Lig-Plot shows Hydrogen bonds as dashed green lines. The residues are
shown as “eyelashes,” which represent interactions other than hydrogen bonding.
Image 9: The Lig-Plot for
flurbiprofen, obtained from PDBSum
database.
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Farhan Ahmed
Protein Modeling Project
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In order to examine the protein-hetero compound interactions in the three-dimensional
structure, the .pdb file was modified in DS Visualizer. First, flurbiprofen was displayed against a
ribbon structure of the protein. Then the interacting amino acids indicated in the LigPlot were
highlighted and changed into the ball and stick structure. The ribbon portion was then removed.
Each amino acid was labeled with its three letter code. The Hydrogen atoms were added to
flurbiprofen, and the hydrogen bonds between the amino acids and flurbiprofen were turned on.
This was done through the H-bond monitor function in DS Visualizer. The results of these steps
can be seen in image 10. Once the Hydrogen bonds were active, the ribbon portion was turned
on and the final rendering of the protein complex was completed, which can be seen in image 11.
Image 10: The interactions of the amino acids with the hetero-compound.
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Farhan Ahmed
Protein Modeling Project
Image 11: The final rendering of the protein-hetero compound.
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Farhan Ahmed
Protein Modeling Project
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Part IV: Protein-Ligand Interactions
H3C
6
O
1
5
OH
2
4
3
F
Image 12: Flurbiprofen (2-(3-fluoro-4-phenyl-phenyl)propanoic acid)
Amino Acid Residue
Trp387
Side Chains- Indole ring
Leu 359
Side Chains- Carbons
Hetero-compound
Atoms
Carbon 1-6 2nd
ring
Methyl
Leu 352
Side Chains- Carbons
2nd ring
Hydrophobic interaction
Gly 526
Side Chains- Hydrogen
…
…
Met 522
Side Chains- Sulfur
2nd ring
Weak non-polar interaction
Ala 527
Side Chain- Carbon
…
Van Der Waals
Val 349
Side Chain- Carbon
Carbon1-6 1st ring
Hydrophobic interaction
Tyr 355
Side chains- Phenol ring
Carboxylate
HBD
Ile523
Side Chains- Carbons
Carbon 1-6 1st ring
Hydrophobic interaction
Arg120
Side Chains- Nitrogen
Carboxylate
HBD
Val116
Side Chains- Carbons
Carboxylate
Dipole induced interaction
Leu531
Side Chains- Carbons
Carbon 1-6 1st ring
Hydrophobic interaction
Tyr 385
Side Chain- Phenol ring
Hydrophobic interaction
Ser 530
Side Chain- CH3OH
Carbon 1-6 2nd
ring
Fluorine
Table 2: Protein-Ligand interactions.
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Nature of Interaction
Hydrophobic interaction
Hydrophobic interaction
HBA
Farhan Ahmed
Protein Modeling Project
Computer 2
Part V: Bibliography
1. Acta Crystallogr D Biol Crystallogr. 2006 Feb; 62(Pt 2):151-6.
2. PDB ID: 2ayl
Gupta K, Selinsky BS, Loll PJ.:
2.0 angstroms structure of prostaglandin H2 synthase-1 reconstituted with a manganese
porphyrin cofactor. Acta crystallographica. Section D, Biological crystallography (2006) 62, pp.
151-6 [PubMed entry 16421446]
3. Laskowski R A (2009). PDBsum new things. Nucleic Acids Res., Database issue, in press.
http://www.ebi.ac.uk/thornton-srv/databases/pdbsum/
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