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. 2024 May 10;29(10):2234.
doi: 10.3390/molecules29102234.

A Combined Molecular Dynamics and Hydropathic INTeraction (HINT) Approach to Investigate Protein Flexibility: The PPARγ Case Study

Federica Agosta  1 , Pietro Cozzini  1
Affiliations

A Combined Molecular Dynamics and Hydropathic INTeraction (HINT) Approach to Investigate Protein Flexibility: The PPARγ Case Study

Federica Agosta et al. Molecules. .

Abstract

Molecular Dynamics (MD) is a computational technique widely used to evaluate a molecular system's thermodynamic properties and conformational behavior over time. In particular, the energy analysis of a protein conformation ensemble produced though MD simulations plays a crucial role in explaining the relationship between protein dynamics and its mechanism of action. In this research work, the HINT (Hydropathic INTeractions) LogP-based scoring function was first used to handle MD trajectories and investigate the molecular basis behind the intricate PPARγ mechanism of activation. The Peroxisome Proliferator-Activated Receptor γ (PPARγ) is an emblematic example of a highly flexible protein due to the extended ω-loop delimiting the active site, and it is responsible for the receptor's ability to bind chemically different compounds. In this work, we focused on the PPARγ complex with Rosiglitazone, a common anti-diabetic compound and analyzed the molecular basis of the flexible ω-loop stabilization effect produced by the Oleic Acid co-binding. The HINT-based analysis of the produced MD trajectories allowed us to account for all of the energetic contributions involved in interconverting between conformational states and describe the intramolecular interactions between the flexible ω-loop and the helix H3 triggered by the allosteric binding mechanism.

Keywords: HINT force field; Molecular Dynamics; PPARγ; conformational analysis; mechanism of action.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
PPARγ active site characteristics. (A) PPARγ active site presents a high volume and is defined between helix3, helix12, and the flexible ω-loop. It can be divided into two regions: the orthosteric site (grey) and the alternate site (orange). (B) PPARγ presents different activation mechanisms. A single compound can bind the orthosteric site with a consequent canonical activation mechanism. The active site high volume allows for the interaction of two or more ligands simultaneously. One of the co-bound ligands is located in the alternate site (PDB ID: 1FM6, 4EM9, and 6MD4).
Figure 2
Figure 2
PPARγ ligand binding mode. Ligand interactions are shown by employing the PLIP plugin in Pymol. Protein is shown in the transparent cartoon, ligand as orange stick, and interaction types are depicted according to PLIP representation: hydrogen bonds as blue lines, salt bridge as dotted yellow lines, and hydrophobic interactions as dotted grey lines. (A) Fatty acids are PPARγ natural ligands (PDB ID: 4EM9) that are able to interact with the Orthosteric binding site through salt bridges with histidine residues and hydrogen bonds with residues in Arm I, while the hydrophobic tail is stabilized through van der Waals interactions. (B) Rosiglitazone (PDB ID: 1FM6) is one of the thiazolidinediones used for type II diabetes treatment and shares the same interaction pathway with fatty acids. (C) Fatty acids can also interact with the alternate site when another ligand is bound to the Orthosteric one (PDB ID: 6MD4). This interaction involves a salt bridge with Arg288 and hydrophobic interactions.
Figure 3
Figure 3
MD analysis. (A) The RMSD analysis of the PPARγ–Rosiglitazone complex (PDB ID: 1FM6) shows the high stability of the system during all simulation times in all three replicas (Run1 in blue, Run2 in orange, and Run3 in grey). (B) The RMSD profile of the PPARγ–Rosiglitazone–Oleic Acid complex (PDB ID:6MD4) in three independent replicas reveals system stability (Run1 in blue, Run2 in orange and Run3 in grey). The protein RMSF profile is shown in (C) (PPARγ–Rosiglitazone system) and (D) (PPARγ–Rosiglitazone–Oleic Acid complex). The red rectangle highlights the flexible regions corresponding to the ω-loop residues (265–276 residues). A second ligand bound to the alternate site produces a significant ω-loop stabilization.
Figure 4
Figure 4
Rosiglitazone and Oleic Acid interactions during the simulation time. Rosiglitazone (A) is stable in both system thanks to h-bond interactions with polar residues (His323, Ser289, and Tyr473). The Oleic Acid (B) adopts two different conformations responsible for its higher RMSD value. These conformations differ for the orientation of the acidic group that can be stabilized through salt bridge interactions with Arg288 (green conformation) or with Lys265 (yellow conformation).
Figure 5
Figure 5
HINT profiles. PPARγ–Rosiglitazone (A) and PPARγ–Rosiglitazone–Oleic Acid complex (B) presents a comparable average total HINT score in all three produced trajectories (Run1, Run2, and Run3 are represented as blue, orange, and grey lines, respectively). Although stable, systems are significantly different for h-bond energy contribution to the total HINT score (PPARγ–Rosiglitazone (C) and PPARγ–Rosiglitazone–Oleic Acid complex (D). This energy contribution is greater in the PPARγ–Rosiglitazone–Oleic Acid complex, suggesting a different intramolecular connection pattern.
Figure 6
Figure 6
HINT profiles. Rosiglitazone–Oleic Acid co-binding effect. (A) PPARγ 3D structure alignment reveals a conformational variation of the ω-loop and helix H3. Based on HINT energy profile analysis, protein 3D structures were extracted from the MD trajectory. PPARγ–Rosiglitazone–Oleic acid complex is represented in green, while the PPARγ–Rosiglitazone complex is in blue. Structural changes are underlined through red arrows. (B) Protein structure reveals an alternative H3 conformation where the Phe282 phenyl ring (green sticks) is projected toward a hydrophobic task (orange residues). (C) ω-loop residues are stabilized thanks to an intensive intramolecular interaction pattern with H3 residues. Hydrogen bonds are shown as blue lines, while electrostatic interactions are shown as orange lines.
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References

    1. Gund P., Halgren T.A., Smith G.M. Annual Reports in Medicinal Chemistry. Academic Press; Cambridge, MA, USA: 1987. Chapter 27 Molecular Modeling as an Aid to Drug Design and Discovery; pp. 269–279. - DOI
    1. Tripathi M.K., Ahmad S., Tyagi R., Dahiya V., Yadav M.K. Computer Aided Drug Design (CADD): From Ligand-Based Methods to Structure-Based Approaches. Elsevier; Amsterdam, The Netherlands: 2022. Fundamentals of molecular modeling in drug design; pp. 125–155. - DOI
    1. Adelusi T.I., Oyedele A.-Q.K., Boyenle I.D., Ogunlana A.T., Adeyemi R.O., Ukachi C.D., Idris M.O., Olaoba O.T., Adedotun I.O., Kolawole O.E., et al. Molecular modeling in drug discovery. Inform. Med. Unlocked. 2022;29:100880. doi: 10.1016/j.imu.2022.100880. - DOI
    1. Hospital A., Goñi J.R., Orozco M., Gelpi J.L. Molecular dynamics simulations: Advances and applications. Adv. Appl. Bioinform. Chem. 2015;8:37–47. doi: 10.2147/aabc.s70333. - DOI - PMC - PubMed
    1. van Gunsteren W.F., Berendsen H.J.C. Computer Simulation of Molecular Dynamics: Methodology, Applications, and Perspectives in Chemistry. Angew. Chem. Int. Ed. 1990;29:992–1023. doi: 10.1002/anie.199009921. - DOI

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