title: Inhibition of human alpha-, beta- and gamma-thrombin by mono-, bis-, tris- and tetra-benzamidine structures: thermodynamic study.
abstract: The inhibitory effect of mono-, bis-, tris- and tetra-benzamidine structures (benzamidine, DAPP, TAPB and TAPP, respectively) on the catalytic properties of human alpha-, beta- and gamma-thrombin (alpha-, beta- and gamma-thrombin, respectively) was investigated (between pH 2.0 and 7.0, I = 0.1 M; T = 37.0 +/- 0.5 degrees C). The affinity of DAPP, TAPB and TAPP for alpha- and beta-thrombin is higher than that found for benzamidine association around neutrality, converging in the acidic pH limb; in contrast, benzamidine, DAPP, TAPB and TAPP show the same value of the association inhibition constant (Ki; M-1) for gamma-thrombin over the whole pH range explored. On lowering the pH from 5.5 to 3.0, the decrease in affinity for benzamidine binding to alpha-, beta- and gamma-thrombin, as well as for DAPP, TAPB and TAPP association to gamma-thrombin reflects the acidic-pK shift, upon inhibitor binding of a single ionizing group. On the other hand, values of Ki for DAPP, TAPB and TAPP binding to alpha- and beta-thrombin appear to be modulated by the acidic-pK shift, upon inhibitor association, of two equivalent proton-binding residues over the same pH range. By considering molecular models of the serine proteinase:inhibitor complexes, the observed binding behaviour of benzamidine, DAPP, TAPB and TAPP to alpha-, beta- and gamma-thrombin has been related to the inferred stereochemistry of the enzyme:inhibitor contact region(s).
authors.affiliation: Department of Pharmaceutical Chemistry and Technology, University of Turin, Italy.
authors.forename: P
authors.initials: P
authors.lastname: Ascenzi
authors.forename: R
authors.initials: R
authors.lastname: Fruttero
authors.forename: G
authors.initials: G
authors.lastname: Amiconi
authors.forename: L
authors.initials: L
authors.lastname: Pugliese
authors.forename: M
authors.initials: M
authors.lastname: Bolognesi
authors.forename: M
authors.initials: M
authors.lastname: Coletta
authors.forename: S
authors.initials: S
authors.lastname: Onesti
authors.forename: M
authors.initials: M
authors.lastname: Guarneri
authors.forename: E
authors.initials: E
authors.lastname: Menegatti
mesh_terms: D000578:Amidines / Q000494:pharmacology; D000595:Amino Acid Sequence; D001550:Benzamidines / Q000737:chemistry / Q000494:pharmacology*; D001665:Binding Sites; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008968:Molecular Conformation; D010879:Piperazines / Q000494:pharmacology; D011485:Protein Binding; D011487:Protein Conformation; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology*; D013816:Thermodynamics; D013917:Thrombin / Q000037:antagonists & inhibitors* / Q000737:chemistry
chemical_list: D000578:Amidines; D001550:Benzamidines; D010879:Piperazines; D015842:Serine Proteinase Inhibitors; C034807:tetra-4-amidinophenoxypropane; C048754:1-(diethylaminopropyl)-4-phenylpiperazine; C074442:1,3-bis(4-amidinophenoxy)-2-(4-amidinophenoxymethyl)ethylpropane; D013917:Thrombin; C032157:benzamidine