Table 1.
Regions of matrix metalloproteinase-2.
Fig 1.
Ribbon representation of the structure of MMP-2 (PDB 1CK7).
Domains identified as follows: collagenase-like 1, orange; collagen binding, green; collagenase-like 2, purple; hinge region, gray; hemopexin, red; Zn2+, red; Ca2+, cyan.
Fig 2.
Ribbon representation of the three-dimensional structure of Ctx.
(A) Random coil, cyan; alpha helix, blue; beta turn, green; residues Gly 24 – Arg 36, pink. Disulfide bridges are shown in sticks. (B) amino acid sequence of Ctx. Disulfide bridges are present between residue pairs (2,19), (5,28), (16,33), (20,35).
Table 2.
Designed peptide fragments of Ctx.
Table 3.
Docking scores and binding energy (∆Eb, kcal/mol) after 100 ns molecular docking of accepted HPEPDOCK poses.
Table 4.
Binding energy (∆Eb) of peptides among docking methods.*.
Fig 3.
Ribbon representation of the central structures of the largest clusters of 500 ns trajectories of peptide – MMP-2 complexes using the re-ranked top HPEPDOCK docking poses.
The orientation of MMP-2 has been optimized to allow for viewing of the peptide-MMP-2 interactions. Peptide ligand regions identified as follows: random coil, cyan; alpha helix, blue; beta sheet, red; beta turn, green. Disulfide bridges are shown in sticks. For MMP-2 receptor: collagenase-1 region, orange; collagenase-2 region, purple; remainder of protein, gray. Abbreviations for MMP-2 domains are from Table 1.
Fig 4.
Residue contribution to ΔEb from MM-PBSA calculations for peptides and MMP-2 complexes obtained from the 500 ns trajectories MD simulations.
(A) Ctx – MMP-2; (B) P75 – MMP-2; (C) P76 – MMP-2; (D) P77 – MMP-2; (E) P78 – MMP-2. Top panel contains residues from MMP-2, bottom panel contains residues from peptides. ΔEcont is the contribution to binding energy for each residue.
Table 5.
Docking scores of accepted HADDOCK poses.
Fig 5.
Representative structures of the largest clusters of 500 ns trajectories of peptide – MMP-2 complexes obtained with the HADDOCK method.
The orientation of MMP-2 has been optimized to allow for viewing of the peptide-MMP-2 interactions. Protein ligand regions identified as follows: random coil, cyan; alpha helix, blue; beta sheet, red; beta turn, green. Disulfide bridges are shown in sticks. For MMP-2 receptor: collagenase-1 region, orange; collagenase-2 region, purple; remainder of protein, gray. Abbreviation for MMP-2 domains are from Table 1.
Fig 6.
Residue-residue decomposition of ΔEb from MMP-BSA calculations of peptides and MMP-2 complexes obtained with the HADDOCK method.
(A) Ctx – MMP-2; (B) P75 – MMP-2; (C) P76 – MMP-2; (D) P77 – MMP-2; (E) P78 – MMP-2. Top values contain residues from MMP-2, bottom values contain residues from peptides. For Ctx, P75, P77, and P78, there were no contributions from MMP-2 residues. ΔEcont is the contribution to binding energy for each residue.
Table 6.
Confidence scores* of accepted poses obtained with the AF2 method.
Fig 7.
Representative structures of the largest clusters of 500 ns trajectories of peptide – MMP-2 complexes obtained with the AF2 method.
Protein ligand regions identified as follows: random coil, cyan; alpha helix, blue; beta sheet, red; beta turn, green. Disulfide bridges are shown in sticks. For MMP-2 receptor: collagenase-1 region, orange; collagenase-2 region, purple; remainder of protein, gray. Abbreviation for MMP-2 domains are from Table 1.
Fig 8.
Residue-residue decomposition of ΔEb from MM-PBSA calculations of peptides − MMP-2 complexes obtained with the AF2 method.
(A) Ctx – MMP-2; (B) P75 – MMP-2; (C) P76 – MMP-2; (D) P77 – MMP-2; (E) P78 – MMP-2. Top values contain residues from MMP-2, bottom values contain residues from peptides. ΔEcont is the contribution to binding energy for each residue.
Table 7.
Change in melting temperature (ΔTm*) due to peptides binding to MMP-2.
Fig 9.
Binding of the peptides to MMP-2 determined by DSF.
Fig 10.
Inhibition of enzymatic activity of MMP-2.
(A) Effects of Ctx at various concentrations on MMP-2 enzymatic activity. Data is presented by mean ± SEM and is not statistically significant. (B) Effects of Ctx, P75, P76, and P78 on MMP-2 enzymatic activity, along with the NNGH positive control. Data is presented by mean and SEM. Compared to the control, the experimental groups are not statistically significant (one-way ANOVA).
Fig 11.
Wound-Healing Assay- U-87 MG cells.
Quantification of cell migration inhibition using the wound-healing assay. Percentage of wound closure was measured using the Wound_Healing_Size_tool_plugin in ImageJ. Results are represented by the mean ± SEM. ***P < 0.0001 (One-way Anova).