Designing allosteric regulators of thrombin. Monosulfated benzofuran dimers selectively interact with Arg173 of exosite 2 to induce inhibition
Research article published in Journal of medicinal chemistry (2012)
Abstract
Earlier, we reported on the design of sulfated benzofuran dimers (SBDs) as allosteric inhibitors of thrombin (Sidhu et al. J. Med. Chem.201154 5522-5531). To identify the site of binding of SBDs, we studied thrombin inhibition in the presence of exosite 1 and 2 ligands. Whereas hirudin peptide and heparin octasaccharide did not affect the IC(50) of thrombin inhibition by a high affinity SBD, the presence of full-length heparin reduced inhibition potency by 4-fold. The presence of γ' fibrinogen peptide, which recognizes Arg93, Arg97, Arg173, Arg175, and other residues, resulted in a loss of affinity that correlated with the ideal Dixon-Webb competitive profile. Replacement of several arginines and lysines of exosite 2 with alanine did not affect thrombin inhibition potency, except for Arg173, which displayed a 22-fold reduction in IC(50). Docking studies suggested a hydrophobic patch around Arg173 as a plausible site of SBD binding to thrombin. The absence of the Arg173-like residue in factor Xa supported the observed selectivity of inhibition by SBDs. Cellular toxicity studies indicated that SBDs are essentially nontoxic to cells at concentrations as high as 250 mg/kg. Overall, the work presents the localization of the SBD binding site, which could lead to allosteric modulators of thrombin that are completely different from all clinically used anticoagulants.
Abstract sourced from PubMed (NCBI) for the cited record. See the original publication for the authoritative version.
Summary
Designing allosteric regulators of thrombin. Monosulfated benzofuran dimers selectively interact with Arg173 of exosite 2 to induce inhibition.
Why This Matters for Hirudotherapy
What the study examined: this medicinal-chemistry study mapped the binding site of synthetic sulfated benzofuran dimers (SBDs) designed as allosteric thrombin inhibitors. Using exosite ligands and alanine substitutions, the authors localized SBD binding to Arg173 of exosite 2 (a 22-fold IC50 reduction when Arg173 was replaced), with docking supporting a hydrophobic patch there; cellular toxicity testing indicated the compounds were essentially nontoxic. Why it matters for the secretome story: the experiments used a hirudin peptide as an exosite-1 reference ligand, and the abstract notes the hirudin peptide and a heparin octasaccharide did not affect SBD inhibition potency, helping confirm the SBDs act at a different site than the leech molecule's classic exosite-1 footprint. Caveat: this is an in vitro and in silico study of engineered synthetic anticoagulants, not leech-derived compounds; hirudin appears here only as a mechanistic probe, so the relevance to leech therapy is indirect and the work is early-stage discovery, not clinical.
Citation
Designing allosteric regulators of thrombin. Monosulfated benzofuran dimers selectively interact with Arg173 of exosite 2 to induce inhibition
Abdel Aziz MH et al. · Journal of medicinal chemistry, 2012
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