FACULTY OF PHARMACY

BPH 2101-MEDICINAL CHEMISTRY

PRACTICAL LAB REPORT
PRACTICAL 2

MOLECULAR MODELLING AND DOCKING ---- COMPUTER

AIDED MOLECULAR MODELLING OF DRUGS

STUDENT’S NAME : Lau Xin Hua

MATRIC NUMBER : QIU-202210-005966

PROGRAMME : Bachelor of Pharmacy

LECTURER’S NAME : Dr. Vijay

DATE OF SUBMISSION: 17/1/2024

Objectives
• To understand and perform docking of ligand.
• To understand and evaluate the bonding of amino acids with functional
groups of protein.

Introduction/Principle
Computer Aided Drug Design (CADD) is a tool used to enhance efficacy and
efficiency of the drug discovery process. There are thousands and millions of various
configurations possible for the binding of ligand and protein such that it would be
exhaustive to study their binding affinity and preferred orientation gradually in
succession. CADD aids scientist to develop drug efficiently by providing the best
possible configurations with well-programmed and curated softwares. Through
evaluation via the docking scores, scientists are able to figure out the most optimal
configurations in a much more quick and convenient manner. In the fields of
structural biology and drug development, molecular docking is a computer technique
that predicts the preferred orientation and binding affinity of a ligand when it interacts
with a target protein. In order to gain insight into the nature and strength of the
protein-ligand interaction, the main objective of molecular docking is to simulate and
predict the binding mode of a ligand within the binding site of a protein. Through
simulating in-silico studies with higher efficiency and efficacy, cost and time in the
drug development process could be saved with ease.

Procedures
The protein target and ligand molecule were chosen. The ligand and protein
preparation were done. The docking site in protein was selected. Protein-ligand
docking was carried out using Autodock docking tool. Different ligand poses were
generated and recorded along with their docking scores.
Results

Fig.1 Docking outcome

Fig. 2 Ligand- Seliciclib visualization in Pymol Fig. 3 Protein – human mutant P53 enzyme

Docking Scores
Discussion

Downloading the files

The applications needed for the practical, namely AutoDock Vina, Pymol and
AutoDock Mgl were downloaded prior to the following steps.
The crystal structure of human mutant P53 enzyme was downloaded from protein
databank as PDB file. The ligand seliciclib was downloaded from PubChem as SDF
file. A folder was created and named ‘docking’ and the files downloaded was
dragged to the new folder. The files were renamed to protein and ligand respectively.

Preparation of docking
The protein PDB file was opened using AutoDock. The water molecules were
removed from the protein and polar hydrogen bonds were added. Kollman charges
were added subsequently. The macromolecule was selected and saved as pdbqt file
in the docking folder. The ligand SDF file was converted into PDB file using Pymol.
The ligand was then set up using AutoDock by clicking Input button then selecting
choose. The ligand was then saved as pdbqt file in the same folder. The ligand and
protein was opened in AutoDock simultaneously in the same screen. Gasteiger
charges were added by selecting protein molecule through grid. Grid box was
opened and the grid dimensions were saved as txt file in the docking folder. A config
file was created in txt form. Information such as ligand, receptor, center x,y,z and
size x,y,z, energy range and exhaustiveness was filled in using the information in the
grid txt file.

Docking
Command prompt was used to run the docking by copying and pasting the path of
the Vina files prior to running the docking procedures. In the next line, the path of
The Scripps Research Institute was copied and pasted. The codes were then added
into the command prompt accurately according to the tutorial video. Docking was ran
and the output was saved in the log.txt file.

Visualization
The protein pdbqt file and the output pdbqt file was opened using Pymol. The
docking scores were visualized in the best nine configurations.
Human P53 protein

The p53 protein plays a crucial role in suppressing tumour by controlling the
processes involved in cell division. It regulates cell division by preventing them from
proliferating rapidly or uncontrollably. The TP53 gene exists in all of the nucleus in
each and every cell in the body in which it is integrated into the deoxyribonucleic acid
(DNA) in the nucleus. When the DNA is damaged by physical trauma such as
ultraviolet radiation, toxic chemicals, and oxidative substances, p53 protein will be
the key in determining the repair of the DNA or self-destructive process (apoptosis)
of the DNA. If the DNA is repairable, p53 protein will be activated and hence initiating
a cascade activating other genes to fix the damaged DNA. If the DNA is
unrepairable, the protein prevents the damaged DNA to continue proliferate and will
induce the cell to undergo apoptosis (Bethesda, 2020).

Inactivation of p53 in cancer cells

The inactivation of p53 tumour suppressor is frequently seen in tumorigenesis.
Mutations in the p53 gene typically produce a stable mutant protein. The
accumulation of this stable mutant protein is thought to be a hallmark of cancerous
cells. Not only do these mutant p53 proteins lose their capacity to inhibit tumour
growth, but they also often pick up additional oncogenic properties that confer growth
and survival advantages to cells (Rivlin et. al, 2011).

Roscovitine and p53 binding

Seliciclib (roscovitine) is a potent cyclin-dependent kinase (CDK) inhibitor. CDKs are
key enzymes regulating cell proliferation through cell checkpoints and transcriptional
events responding to extracellular and intracellular signals (Ding et. al, 2020).
Roscovitine can inhibit CDK by directly binding to the ATP binding sites of the CDK.
Since p53 can be a direct downstream kinase substrate for CDKs (Morris et. al,
2001), roscovitine and p53 binding are plausible for the comparation of the docking
scores in the aspect of medicinal chemistry and is plausible to be investigated
further. Moreover, roscovitine is also able to target p53 pathway and induce
apoptosis of cancerous cells (Dey et. al, 2008). Roscovitine has the ability to restore
mutant p53 by refolding the protein and prevent its aggregation (Wang et. al, 2023).
Biological activity of roscovitine
Seliciclib/roscovitine belongs to the purine family, which all shares the basic ring
structure which is composed of pyrimidine and imidazole. Important components in
the purine family include adenine, guanine and ATP. It functions by binding with the
amino acids that line up the ATP-binding pocket of the CDK catalytic domain and
competes with ATP for binding at the ATP-binding site of CDKs. In the case of CDK2
from the CDK family, two hydrogen bonds are involved in most of the interaction
between CDK2 and roscovitine. This interaction is formed between N6 and N7 of the
purine and backbone atoms of leucine 83. Additionally, O1 and a water molecule
establish a weak hydrogen bond, and roscovitine's benzyl ring faces away from
the ATP-binding pocket. Hence, ATP cannot bind to the kinase and a catalytic
reaction cannot be performed (Cicenas et. al, 2015). Thus, cellular proliferation is
unable to occur without ATP production. Furthermore, one hydrogen bond donor
with hydrophobic interaction of roscovitine with phenylalanine 82, isoleucine 10,
leucine 134, and alanine 31 are also essential for its biological activity to prevent
ATP binding (El-Sattar et. al, 2021)

Docking Scores Interpretation

The term affinity in the unit kcal/mol indicates how tightly a ligand and a protein bind.
The distance from the best modes is measured in the unit of rmsd u.b and rmsd l.d.
which indicates the similarity of the other modes compared to best mode in terms of
the position of the atoms. Lower values of rmsd also indicate higher similarity of the
modes compared to best mode. The best mode has an affinity of -5.8 kcal/mol since
it has a lower/ more negative energy value, meaning that it binds more strongly.

Conclusion
Docking between the mutant p53 protein and seliciclib/roscovitine was carried out
successfully. The best mode was the mode with -5.8 kcal/mol energy since it has the
lowest value.

References

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