Biochemical mechanism of action of a diketopiperazine inactivator of plasminogen activator inhibitor-1.

Abstract

XR5118 [(3 Z,6 Z )-6-benzylidine-3-(5-(2-dimethylaminoethyl-thio-))-2-(thienyl)methylene-2,5-dipiperazinedione hydrochloride] can inactivate the anti-proteolytic activity of the serpin plasminogen activator inhibitor-1 (PAI-1), a potential therapeutic target in cancer and cardiovascular diseases. Serpins inhibit their target proteases by the P(1) residue of their reactive centre loop (RCL) forming an ester bond with the active-site serine residue of the protease, followed by insertion of the RCL into the serpin's large central beta-sheet A. In the present study, we show that the RCL of XR5118-inactivated PAI-1 is inert to reaction with its target proteases and has a decreased susceptibility to non-target proteases, in spite of a generally increased proteolytic susceptibility of specific peptide bonds elsewhere in PAI-1. The properties of XR5118-inactivated PAI-1 were different from those of the so-called latent form of PAI-1. Alanine substitution of several individual residues decreased the susceptibility of PAI-1 to XR5118. The localization of these residues in the three-dimensional structure of PAI-1 suggested that the XR5118-induced inactivating conformational change requires mobility of alpha-helix F, situated above beta-sheet A, and is in agreement with the hypothesis that XR5118 binds laterally to beta-sheet A. These results improve our understanding of the unique conformational flexibility of serpins and the biochemical basis for using PAI-1 as a therapeutic target.

• Authors: Anja P Einholm, Katrine E Pedersen, Troels Wind, Paulina Kulig, Michael T Overgaard, Jan K Jensen, Julie S Bødker, Anni Christensen, Peter Charlton, Peter A Andreasen
• Journal: The Biochemical journal (Biochem J) Vol. 373 Issue Pt 3 Pg. 723-32 (Aug 01 2003) ISSN: 0264-6021 [Print] England
• PMID: 12723974 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Piperazines
• Plasminogen Activator Inhibitor 1
• Vitronectin
• XR 5118

Topics

• Humans
• Hydrolysis
• Models, Molecular
• Mutagenesis, Site-Directed
• Piperazines (pharmacology)
• Plasminogen Activator Inhibitor 1 (chemistry, genetics)
• Vitronectin (metabolism)