title: Binding of hirudin to human alpha, beta and gamma-thrombin. A comparative kinetic and thermodynamic study.
abstract: Thermodynamic parameters for the binding of hirudin to human alpha, beta and gamma-thrombin have been determined between pH 5.0 and 9.0, and from 10 degrees C to 40 degrees C; kinetic data for the association and dissociation of the proteinase-inhibitor complex were obtained at pH 7.5 and 21 degrees C. These results have been analysed in parallel with the inhibitor-binding properties of human alpha, beta and gamma-thrombin for the bovine basic pancreatic trypsin inhibitor (Kunitz-type inhibitor; BPTI). For the purpose of an homogeneous comparison, values of the apparent association equilibrium constant for BPTI binding to human gamma-thrombin have been determined between pH 5.0 and 9.0, at 21 degrees C. The different binding behaviour of hirudin and BPTI with respect to human alpha, beta and gamma-thrombin has been related to the inferred stereochemistry of the proteinase-inhibitor contact regions. In particular, whereas the beta and gamma-loops play an appreciable role in the stabilization of the enzyme-hirudin complexes, they contribute to impairment of the adduct formation for the proteinase/BPTI system.
authors.affiliation: Department of Pharmaceutical Chemistry and Technology, University of Turin, Italy.
authors.forename: P
authors.initials: P
authors.lastname: Ascenzi
authors.forename: G
authors.initials: G
authors.lastname: Amiconi
authors.forename: M
authors.initials: M
authors.lastname: Coletta
authors.forename: G
authors.initials: G
authors.lastname: Lupidi
authors.forename: E
authors.initials: E
authors.lastname: Menegatti
authors.forename: S
authors.initials: S
authors.lastname: Onesti
authors.forename: M
authors.initials: M
authors.lastname: Bolognesi
mesh_terms: D007611:Aprotinin / Q000737:chemistry; D006629:Hirudins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D011485:Protein Binding; D011487:Protein Conformation; D013816:Thermodynamics; D013917:Thrombin / Q000737:chemistry / Q000378:metabolism*
chemical_list: D006629:Hirudins; D007611:Aprotinin; D013917:Thrombin