title: Dilutional control of prothrombin activation at physiologically relevant shear rates.
abstract: The generation of proteolyzed prothrombin species by preassembled prothrombinase in phospholipid-coated glass capillaries was studied at physiologic shear rates (100-1000 s(-1)). The concentration of active thrombin species (α-thrombin and meizothrombin) reaches a steady state, which varies inversely with shear rate. When corrected for shear rate, steady-state levels of active thrombin species exhibit no variation and a Michaelis-Menten analysis reveals that chemistry of this reaction is invariant between open and closed systems; collectively, these data imply that variations with shear rate arise from dilutional effects. Significantly, the major products observed include nonreactive species arising from the loss of prothrombin's phospholipid binding domain (des F1 species). A numerical model developed to investigate the spatial and temporal distribution of active thrombin species within the capillary reasonably approximates the observed output of total thrombin species at different shears; it also predicts concentrations of active thrombin species in the wall region sufficient to account for observed levels of des FI species. The predominant feedback formation of nonreactive species and high levels of the primarily anticoagulant intermediate meizothrombin (∼40% of total active thrombin species) may provide a mechanism to prevent thrombus propagation downstream of a site of thrombosis or hemorrhage.
authors.affiliation: Department of Biochemistry, University of Vermont, College of Medicine, Colchester, Vermont.
authors.forename: Laura M
authors.initials: LM
authors.lastname: Haynes
authors.affiliation: Mechanical Engineering Department, University of Vermont, College of Engineering and Mathematical Sciences, Burlington, Vermont.
authors.forename: Yves C
authors.initials: YC
authors.lastname: Dubief
authors.affiliation: Department of Biochemistry, University of Vermont, College of Medicine, Colchester, Vermont.
authors.forename: Thomas
authors.initials: T
authors.lastname: Orfeo
authors.affiliation: Department of Biochemistry, University of Vermont, College of Medicine, Colchester, Vermont. Electronic address: kenneth.mann@uvm.edu.
authors.forename: Kenneth G
authors.initials: KG
authors.lastname: Mann
mesh_terms: D000818:Animals; D002417:Cattle; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000378:metabolism; D018056:Hemorheology; D006801:Humans; D007700:Kinetics; D008954:Models, Biological; D017434:Protein Structure, Tertiary; D011516:Prothrombin / Q000737:chemistry / Q000378:metabolism*; D013314:Stress, Mechanical; D013917:Thrombin / Q000378:metabolism; D013925:Thromboplastin / Q000378:metabolism
chemical_list: D004792:Enzyme Precursors; D011516:Prothrombin; D013925:Thromboplastin; C012007:prethrombins; D013917:Thrombin; C026812:meizothrombin