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Characterization of the Annonaceous acetogenin, annonacinone, a natural product inhibitor of plasminogen activator inhibitor-1

View Article: PubMed Central - PubMed

ABSTRACT

Plasminogen activator inhibitor-1 (PAI-1) is the main inhibitor of the tissue type and urokinase type plasminogen activators. High levels of PAI-1 are correlated with an increased risk of thrombotic events and several other pathologies. Despite several compounds with in vitro activity being developed, none of them are currently in clinical use. In this study, we evaluated a novel PAI-1 inhibitor, annonacinone, a natural product from the Annonaceous acetogenins group. Annonacinone was identified in a chromogenic screening assay and was more potent than tiplaxtinin. Annonacinone showed high potency ex vivo on thromboelastography and was able to potentiate the thrombolytic effect of tPA in vivo in a murine model. SDS-PAGE showed that annonacinone inhibited formation of PAI-1/tPA complex via enhancement of the substrate pathway. Mutagenesis and molecular dynamics allowed us to identify annonacinone binding site close to helix D and E and β-sheets 2A.

No MeSH data available.


Depiction of a putative binding mode of annonacinone against the active form of PAI-1.(A) Overall view of the complex, with the solvent accessible surface depicted as semi-transparent cyan when polar, and purple when nonpolar. (B) Detailed view of the binding site. Cyan lines indicate hydrogen bonds. The name of residues identified by site directed mutagenesis is printed in yellow.
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f5: Depiction of a putative binding mode of annonacinone against the active form of PAI-1.(A) Overall view of the complex, with the solvent accessible surface depicted as semi-transparent cyan when polar, and purple when nonpolar. (B) Detailed view of the binding site. Cyan lines indicate hydrogen bonds. The name of residues identified by site directed mutagenesis is printed in yellow.

Mentions: Overall examination of the molecular dynamics trajectory of the complex reveals that while annonacinone and PAI-1 remain bound to each other, the long aliphatic chain is the subject of large amplitude fluctuations from its position obtained by docking, and eventually seems to adhere to the hydrophobic surface of PAI-1 in the neighborhood of the vitronectin binding site (Fig. 5A) notably to the side chain of Phe114 as well as the non-terminal, hydrophobic part of that of Arg115. Investigation and comparison of the volume of the binding site in the presence and absence of annonacinone showed a lower average value when the ligand is bound (see Supplementary Fig. S5). At the same time, movement of the reactive loop was found of lower amplitude when the binding site is occupied by annonacinone (see Supplementary Fig. S6). A closer look at the reference ligand binding pocket shows that the α,β-unsaturated lactone head of annonacinone penetrates more deeply into it during the course of the simulation, progressively establishing several hydrogen bonds, notably with residues Asp95 and Arg118 (Fig. 5B), which were also identified by site-directed mutagenesis. Although the hydrogen bond is not visible in Fig. 5B, Lys122 residue participated a significant proportion of the time, as well as backbone atoms belonging to Ser119 and Val121.


Characterization of the Annonaceous acetogenin, annonacinone, a natural product inhibitor of plasminogen activator inhibitor-1
Depiction of a putative binding mode of annonacinone against the active form of PAI-1.(A) Overall view of the complex, with the solvent accessible surface depicted as semi-transparent cyan when polar, and purple when nonpolar. (B) Detailed view of the binding site. Cyan lines indicate hydrogen bonds. The name of residues identified by site directed mutagenesis is printed in yellow.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5120274&req=5

f5: Depiction of a putative binding mode of annonacinone against the active form of PAI-1.(A) Overall view of the complex, with the solvent accessible surface depicted as semi-transparent cyan when polar, and purple when nonpolar. (B) Detailed view of the binding site. Cyan lines indicate hydrogen bonds. The name of residues identified by site directed mutagenesis is printed in yellow.
Mentions: Overall examination of the molecular dynamics trajectory of the complex reveals that while annonacinone and PAI-1 remain bound to each other, the long aliphatic chain is the subject of large amplitude fluctuations from its position obtained by docking, and eventually seems to adhere to the hydrophobic surface of PAI-1 in the neighborhood of the vitronectin binding site (Fig. 5A) notably to the side chain of Phe114 as well as the non-terminal, hydrophobic part of that of Arg115. Investigation and comparison of the volume of the binding site in the presence and absence of annonacinone showed a lower average value when the ligand is bound (see Supplementary Fig. S5). At the same time, movement of the reactive loop was found of lower amplitude when the binding site is occupied by annonacinone (see Supplementary Fig. S6). A closer look at the reference ligand binding pocket shows that the α,β-unsaturated lactone head of annonacinone penetrates more deeply into it during the course of the simulation, progressively establishing several hydrogen bonds, notably with residues Asp95 and Arg118 (Fig. 5B), which were also identified by site-directed mutagenesis. Although the hydrogen bond is not visible in Fig. 5B, Lys122 residue participated a significant proportion of the time, as well as backbone atoms belonging to Ser119 and Val121.

View Article: PubMed Central - PubMed

ABSTRACT

Plasminogen activator inhibitor-1 (PAI-1) is the main inhibitor of the tissue type and urokinase type plasminogen activators. High levels of PAI-1 are correlated with an increased risk of thrombotic events and several other pathologies. Despite several compounds with in vitro activity being developed, none of them are currently in clinical use. In this study, we evaluated a novel PAI-1 inhibitor, annonacinone, a natural product from the Annonaceous acetogenins group. Annonacinone was identified in a chromogenic screening assay and was more potent than tiplaxtinin. Annonacinone showed high potency ex vivo on thromboelastography and was able to potentiate the thrombolytic effect of tPA in vivo in a murine model. SDS-PAGE showed that annonacinone inhibited formation of PAI-1/tPA complex via enhancement of the substrate pathway. Mutagenesis and molecular dynamics allowed us to identify annonacinone binding site close to helix D and E and β-sheets 2A.

No MeSH data available.