Molecular Dynamics Simulations of Matrix Metalloproteinase 13 and the Analysis of the Specificity Loop and the S1'-Site

Abstract

The specificity loop of Matrix Metalloproteinases (MMPs) is known to regulate recognition of their substrates, and the S1'-site surrounded by the loop is a unique place to address the selectivity of ligands toward each MMP. Molecular dynamics (MD) simulations of apo-MMP-13 and its complex forms with various ligands were conducted to identify the role of the specificity loop for the ligand binding to MMP-13. The MD simulations showed the dual role of T247 as a hydrogen bond donor to the ligand, as well as a contributor to the formation of the van der Waal surface area, with T245 and K249 on the S1'-site. The hydrophobic surface area mediated by T247 blocks the access of water molecules to the S1'-site of MMP-13 and stabilizes the ligand in the site. The F252 residue is flexible in order to search for the optimum location in the S1'-site of the apo-MMP-13, but once a ligand binds to the S1'-site, it can form offset π-π or edge-to-π stacking interactions with the ligand. Lastly, H222 and Y244 provide the offset π-π and π-CH(Cβ) interactions on each side of the phenyl ring of the ligand, and this sandwiched interaction could be critical for the ligand binding to MMP-13.

Keywords: MD simulations; S1′−site; matrix metalloproteinase 13; selective MMP inhibitors; specificity loop; π−CH(Cβ) interactions.

Figure 1

MD simulations of the apo−MMP−13 structure: (A) The specificity loop and S1′−site of MMP-13; Residues in the specificity loop and S1′−site are light blue, and residues in the Met−turn are yellow; (B) RMSD of MD simulations of apo−MMP-13; (C) RMSF of all residues in MD run 1 (blue), run 1 (48–100 ns, red), and run 2 (black); (D) Dihedral angle analysis of F252: (top) MD run 2 (0–100 ns) and (bottom) MD run 1 (48–100 ns); (E) The specificity loops of MMP−13 from MD simulations; the green loop corresponds to the X-ray crystal structure (PDB code: 4L19).; (F) The conformation of T247 and F252 from the MD simulations of MMP-13; the green loop corresponds to the X-ray crystal structure (PDB code: 4L19).; (G) The χ−angle histogram of T247 from the MD simulations (20–100 ns) and the χ−angle of T247 in the X-ray co−crystal structure.

Figure 2

MD simulations of the MMP−13 − 1UA, C1, or C2 complex: (A) The structures of 1UA, C1, and C2; (B) The open conformation of the specificity loop of MMP−13; (C) The H−bond histogram analysis of MMP−13 − 1UA complex; simulation 1 (binding pose 1) and simulation 2 (binding pose 2). The yellow line represents the presence of the H−bond interaction at each time point.; LIG_O and LIG_NH stand for carbonyl oxygen and NH of 1UA, and T245_O and T247_OH for oxygen of the T245 amide backbone and OH of the T247 residue, respectively; (D) Two different conformations of T247 depending on the φ angle of Y246; (E) The binding pose 1 of 1UA; (F) The binding pose 2 of 1UA; (G) The per−residue energy decomposition analyses of MMP−13 − 1UA complex; the binding pose 1 (left) and the binding pose 2 (right).

Figure 3

Analysis of MD simulations of MMP−13 − C1 complex: (A) The per−residue energy decomposition analyses of MMP−13 − C1 complex; (B) The H−bond histogram analysis of MMP−13 − C1 complex; the yellow line represents the presence of the H−bond interaction at each time point. Only the H−bond interactions in the Zn−binding site are present.; LIG_O1, O2, and O3 stand for oxygen of C1, and LIG_N1H and N2H for hydrogen on the amide unit of C1 (left panel). L185_NH/I243_NH/T245_NH and G183_O/P242_O stand for hydrogen and oxygen on the amide backbone of each amino acid, respectively.

Figure 4

Analysis of MD simulations of MMP−13 complexed with various ligands occupying S1′-site: (A) Structures of ligands used for MD simulations; a–f are scaffolds of ligands, and the complete 2−D structures of ligands are present in Table S4.; (B) A data plot of pIC₅₀ vs. binding affinities calculated from MM/GBSA; (C) RMSD of MD simulations of MMP−13 complexed with C99, C100, C101, and C102; (D) The offset π−π stacking interactions of benzene and fluorobenzene obtained from QM calculations; (E) The F252 energy decomposition analyses of MMP−13 complexed with C99−C110 via MM/GBSA calculations; (F) The shift of C103 (green) in the S1′−site due to additional H−bond interactions.

Figure 5

The specificity loop and S1′−site of MMP−13 complexed with ligands: (A–C) The NMR structure of MMP−13 (pdb code: 1FM1) and the X-ray co-crystal structures of MMP−13 (pdb codes: 3TVC and 2PJT), respectively, which shows the flexible conformation of T144/T247 (yellow). The part of each ligand in the S1′-site is shown in green, and each ligand code is labeled; (D,E) The edge−to−face and offset π−π stacking interactions between F252 and ligands in the S1′−site of MMP−13, respectively; (F) The S1′−site closed by the hydrophobic surface made by G237, T245, T247, and K249 in the specificity loop; (G) The open state of S1′−site; (H) The electrostatic potential surface of the offset π–π stacking interaction of benzene and fluorobenzene; (I) The sandwiched π−π and π−CH (Cβ) interactions by the ligand, H222, and Y244, which could be critical for the ligand binding to MMP−13.