title: Heparin modulation of human plasma kallikrein on different substrates and inhibitors.
abstract: The interplay of different proteases and glycosaminoglycans is able to modulate the activity of the enzymes and to affect their structures. Human plasma kallikrein (huPK) is a proteolytic enzyme involved in intrinsic blood clotting, the kallikrein-kinin system and fibrinolysis. We investigated the effect of heparin on the action, inhibition and secondary structure of huPK. The catalytic efficiency for the hydrolysis of substrates by huPK was determined by Michaelis-Menten kinetic plots: 5.12x10(4) M-1 s-1 for acetyl-Phe-Arg-p-nitroanilide, 1.40x10(5) M-1 s-1 for H-D-Pro-Phe-Arg-p-nitroanilide, 2.25x10(4) M-1 s-1 for Abz-Gly-Phe-Ser-Pro-Phe-Arg-Ser-Ser-Arg-Gln-EDDnp, 4.24x10(2)M-1 s-1 for factor XII and 5.58x10(2) M-1 s-1 for plasminogen. Heparin reduced the hydrolysis of synthetic substrates (by 2.0-fold), but enhanced factor XII and plasminogen hydrolysis (7.7- and 1.4-fold, respectively). The second-order rate constants for inhibition of huPK by antithrombin and C1-inhibitor were 2.40x10(2) M-1 s-1 and 1.70x10(4) M-1 s-1, respectively. Heparin improved the inhibition of huPK by these inhibitors (3.4- and 1.4-fold). Despite the fact that huPK was able to bind to a heparin-Sepharose matrix, its secondary structure was not modified by heparin, as monitored by circular dichroism. These actions may have a function in the control or maintenance of some pathophysiological processes in which huPK participates.
authors.affiliation: Departamento de Bioquímica, Universidade Federal de São Paulo, 04044-020 São Paulo, SP, Brazil.
authors.forename: Andrezza J
authors.initials: AJ
authors.lastname: Gozzo
authors.forename: Viviane A
authors.initials: VA
authors.lastname: Nunes
authors.forename: Ilana
authors.initials: I
authors.lastname: Cruz-Silva
authors.forename: Adriana K
authors.initials: AK
authors.lastname: Carmona
authors.forename: Helena B
authors.initials: HB
authors.lastname: Nader
authors.forename: Adelaide
authors.initials: A
authors.lastname: Faljoni-Alario
authors.forename: Misako U
authors.initials: MU
authors.lastname: Sampaio
authors.forename: Mariana S
authors.initials: MS
authors.lastname: Araújo
mesh_terms: D000991:Antithrombins / Q000494:pharmacology; D002384:Catalysis; D050718:Complement C1 Inhibitor Protein / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005174:Factor XII / Q000187:drug effects / Q000378:metabolism; D006493:Heparin / Q000494:pharmacology*; D006801:Humans; D006868:Hydrolysis; D010455:Peptides / Q000187:drug effects / Q000378:metabolism*; D020842:Plasma Kallikrein / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism*; D010958:Plasminogen / Q000187:drug effects / Q000378:metabolism; D017433:Protein Structure, Secondary; D013997:Time Factors; D000515:alpha 1-Antitrypsin / Q000494:pharmacology
chemical_list: D000991:Antithrombins; D050718:Complement C1 Inhibitor Protein; D004791:Enzyme Inhibitors; D010455:Peptides; D000515:alpha 1-Antitrypsin; D005174:Factor XII; D010958:Plasminogen; D006493:Heparin; D020842:Plasma Kallikrein