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Protease-activated receptor 4-like peptides bind to thrombin through an optimized interaction with the enzyme active
David B Cleary1, Toni A Trumbo, Muriel C Maurer
1Department of Chemistry, University of Louisville, 2320 South Brook Street, Louisville, KY 40292, USA.
Abstract:
Protease-activated receptor 4 (PAR4) is cleaved by thrombin at the R47-G48 peptide bond. Unlike PAR1, PAR4 does not contain a sequence readily predicted to interact with thrombin anion binding exosite-I. HPLC kinetic results on hydrolysis of PAR4 peptides (38-51 and 38-62) reveal that extending the sequence from the active site toward the exosite does not promote further binding interactions with thrombin. One-dimensional-proton line-broadening NMR indicates that the amino acids occupying the P(4)-P(1) positions of PAR4 (38-47), 44PAPR(47), come into direct contact with the thrombin surface. Less contact arises from the Leu43 at the P(5) position. Two-dimensional total correlation spectroscopy and two-dimensional transferred nuclear Overhauser effect spectroscropy studies on this complex reveal that Leu43 is flexible and can exhibit two conformational states. The binding mode observed for PAR4 peptides is similar to that of PAR1 peptides. PAR4 takes advantage of a distinctive sequence to optimize its interactions with the thrombin active site surface.