1. Unraveling the mechanism of CETP inhibition through Molecular Dynamics simulations: Insights for rational drug discovery
Revathi.S
BT12S020

2. Introduction

3. Coronary artery diseases and CETP
Atherosclerosis - leading cause of mortality in industrialized nations.
Recent studies have shown that Cholesteryl Ester Transfer Protein (CETP), which modulates the neutral lipid profile in the body between HDL and LDL, is an effective target.
Koivuniemi, A., T. Vuorela, et al. (2012). PLoS Computational Biology 8(1).

4. Cholesteryl ester transfer protein
Boomerang shaped protein molecule with a hydrophobic tunnel.
Transfers lipids between High Density Lipoprotein (HDL) and Low Density Lipoprotein (LDL)
Inhibiting CETP caused reduction of plaques in the arterial walls of rabbits.
Qiu, X., A. Mistry, et al. (2007) Nature structural & molecular biology 14(2):
Zhang, L., F. Yan, et al. (2012) Nature chemical biology 8(4):

5. Currently available inhibitors of CETP
Objective
To study the mechanism of CETP inhibition by the
3,5-bis(trifluoromethy)phenyl derivatives

6. Methodology

7. Molecular dynamics Simulations
Biomolecular simulations – Computational microscope for molecular biology.
Molecular Dynamics is based on Newton’s laws of motion
-𝜵E=Fi
Non-bonded
Bonded
Computer Simulations of Liquids Allen and Tilldeseley

8. Why do we need HPCE for MD?
System under study:
Protein of 7500 atoms protein
522 lipid atoms
81 atoms of inhibitor (variable)
268 Na+ 279 Cl- ions
atoms from water
Total of over atoms

9. Software used: NAMD2.9
System scales up to 11ns/day with 256 CPUs in VIRGO

10. Flow Chart Ligand structure optimization by Gaussian 3.0
Docking of the inhibitors to CETP by AutoDock4
Molecular Dynamics Simulations by NAMD2.9 with CHARMM36 forcefield
Visualization and Analysis using VMD and GROMACS

11. Results
RMSD as a function of time reveals that the systems have stabilized.
RMSF results show that the N-terminal distal region of the inhibitor bound CETP has higher atomic fluctuations in comparison to the substrate-bound complex.

12. Dynamic Cross Correlation Analysis
Anti-correlation
Positive -correlation
Substrate bound CETP
Torcetrapib bound CETP
Anacetrapib bound CETP
Evacetrapib bound CETP
Inhibitor binding increases the anti-correlation between the binding site and distal regions of CETP
Grant, Rodrigues, ElSawy, McCammon, Caves, (2006) Bioinformatics 22,  

13. Principal Component Analysis
Inhibitor bound complexes are more dynamic than the substrate bound complex
The above plot reveals that the first 20 Eigen vectors are required to describe 80% of the overall dynamics of the protein.
Pronk, S., S. Páll, et al. (2013). Bioinformatics (Oxford, England) 29(7):

14. Protein Dynamics from PCA : PC1

15. Protein Dynamics from PCA : PC2

16. PCA : Porcupine plots
PC1
PC2
Substrate bound
Inhibitor bound
Inhibitor bound CETP shows greater extent of twisting in PC2
Humphrey, W., Dalke, A. and Schulten, K., "VMD - Visual Molecular Dynamics", J. Molec. Graphics, 1996, vol. 14, pp

17. APBS Analysis Substrate-bound 0 ns 50 ns 100 ns 150 ns
The N-terminal distal region of both the complexes become predominantly electropositive over time .
The inhibitor bound complex becomes relatively more electropositive.
Torcetrapib-bound
Anacetrapib-bound
Evacetrapib-bound
Baker NA, Sept D, Joseph S, Holst MJ, McCammon JA. Electrostatics of nanosystems: application to microtubules and the ribosome. Proc. Natl. Acad. Sci. USA 98,

18. In conclusion..
Inhibitor binding affects the atomic fluctuations at the distal regions of CETP.
Inhibitor binding increases the anti-correlation between the binding site and distal regions of CETP.
PCA reveals increased dynamics of the inhibitor bound complexes.
Dynamic Cross Correlation Analysis and PCA are indicative of differential twists in the inhibitor-bound systems.
This twisting results in the progressive exposure of electropositive residues in the N-terminal distal region.
Improved electro-positivity at the HDL –sensing N-terminus supports the existing hypothesis of high affinity of inhibitor bound complexes towards HDL having electronegative surface thereby resulting in the formation of non-productive CETP-HDL complex.*
*Clark RW, Ruggeri RB, Cunningham D, Bamberger MJ J Lipid Res 2006;47:537–552

19. Acknowledgements THANK YOU…
Principal Investigator : Dr.Sanjib Senapati
Members of the Computational Biophysics Lab, IITM
HPCE Team:
Mr. V. Ravichandran
Mrs.P.Gayathri
THANK YOU…