title: Reversible modulation of human factor Xa activity with phosphonate esters: media effects.
abstract: Enantiomers of 4-nitrophenyl 4-X-phenacyl methylphosphonate esters (X = H, PMN; CH3 and CH3O) inactivate human factor Xa with rate constants 8-86 M(-1)s(-1) at pH 6.75 in 0.025 M Hepes buffer, 0.15 M NaCl and 2 mM CaCl2 at 7.0+/-0.1 degrees C. The stereoselectivity of the inactivation of factor Xa is 2-10 and favors the levorotatory enantiomers. The pH-dependence of inactivation of factor Xa by (-)-PMN is sigmoidal and consistent with the participation of a catalytic residue with a pKa of 6.2+/-0.1. Factor Xa reactivates from its phosphonyl adducts through a self-catalyzed intramolecular reaction, which is much influenced by the presence of phospholipids. The rate of reactivation in the absence of phospholipids is not pH dependent at pH <9, but it increases very much at pH >9. In the presence of phospholipids, the pH dependence of the rate constant for reactivation is sigmoidal in the pH 6.5-10.3 range and levels off at pH >9 indicating that the enzyme catalyzes its reactivation. The kinetic pKa for the recovery of factor Xa from its adducts with the PMNs is in the range of 6.7-8.1 and is consistent with the participation of the catalytic His57 in the reactivation process.
authors.affiliation: Department of Chemistry, The Catholic University of America, Washington, DC 20064, USA.
authors.forename: E J
authors.initials: EJ
authors.lastname: Enyedy
authors.forename: I M
authors.initials: IM
authors.lastname: Kovach
mesh_terms: D001665:Binding Sites; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology; D004951:Esterification; D015943:Factor Va / Q000494:pharmacology; D015951:Factor Xa / Q000378:metabolism*; D065427:Factor Xa Inhibitors; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008024:Ligands; D008051:Lipid Bilayers / Q000494:pharmacology; D008956:Models, Chemical; D063065:Organophosphonates / Q000737:chemistry / Q000494:pharmacology*; D010718:Phosphatidylserines / Q000494:pharmacology; D010743:Phospholipids / Q000494:pharmacology; D012694:Serine / Q000737:chemistry; D013237:Stereoisomerism
chemical_list: C402786:1,2-dicaproylphosphatidylserine; D004791:Enzyme Inhibitors; D065427:Factor Xa Inhibitors; D008024:Ligands; D008051:Lipid Bilayers; D063065:Organophosphonates; D010718:Phosphatidylserines; D010743:Phospholipids; D012694:Serine; D015943:Factor Va; D015951:Factor Xa