Plasmin-Mediated Activation and Inactivation of Thrombin-Activatable Fibrinolysis Inhibitor

Abstract

Activated thrombin-activatable fibrinolysis inhibitor (TAFIa) attenuates the fibrin cofactor function of tissue-type plasminogen activator-mediated plasmin formation and subsequently fibrin degradation. In the present study, we focused on the role of plasmin in the regulation of TAFIa activity. Upon incubation with plasmin, TAFIa activity was generated, which was unstable at 37 degrees C. Analysis of the cleavage pattern showed that TAFI was cleaved at Arg(92), releasing the activation peptide from the 35.8-kDa catalytic domain. The presence of the 35.8-kDa fragment paralleled the time course of generation and loss of TAFIa activity. This suggested that, in the presence of plasmin, TAFIa is probably inactivated by proteolysis rather than by conformational instability. TAFI was also cleaved at Arg(302), Lys(327), and Arg(330), resulting in a approximately 44.3-kDa fragment and several smaller fragments. The 44.3-kDa fragment is no longer activatable since it lacks part of the catalytic center. We concluded that plasmin can cleave at several sites in TAFI and that this contributes to the regulation of TAFI and TAFIa.

Generation and Characterization of a Highly Stable Form of Activated Thrombin-activable Fibrinolysis Inhibitor

“…This is consistent with the observation that plasmin had no influence on CPB activity at the concentrations tested and that CPB is a very good attenuator of lysis. Furthermore, the fact that plasmin could both activate TAFI-CPB-(293-401) and inactivate TAFIa-CPB-(293-401) substantiated our earlier observation that plasmin inactivates TAFIa via proteolysis (12).…”

“…The latter is cleaved further at several C-terminal sites, resulting in a polypeptide of 25-kDa and some smaller fragments. Some C-terminal cleavages can also occur prior to removal of the activation peptide, resulting in an ϳ44-kDa fragment that is no longer activable because it lacks critical amino acids involved in catalysis (12). The plasmin cleavage patterns of TAFI and TAFI-CPB-(293-333) seemed similar to that of TAFI purified from plasma (12) as assessed by SDS-PAGE analysis (Fig.…”

“…TAFIa is inactivated by conformational instability, after which it becomes more susceptible to proteolytic degradation by thrombin (4,(7)(8)(9)(10)(11). Plasmin can inactivate TAFIa by proteolysis of the catalytic domain (12).…”

Thrombin Activatable Fibrinolysis Inhibitor Activation Peptide Shows Association With All Major Subtypes of Ischemic Stroke and With TAFI Gene Variation

“…It can be activated by trypsin-like enzymes such as thrombin, plasmin, or the thrombin/thrombomodulin complex. [3][4][5] On activation of intact TAFI, the activation peptide (AP) (Phe1-Arg92; 20 kDa) is released from the catalytic domain (TAFIa) (Ala93-Val401; 36 kDa). 4 Both in vitro and in vivo experiments show that TAFIa retards fibrinolysis.…”

“…[3][4][5] On activation of intact TAFI, the activation peptide (AP) (Phe1-Arg92; 20 kDa) is released from the catalytic domain (TAFIa) (Ala93-Val401; 36 kDa). 4 Both in vitro and in vivo experiments show that TAFIa retards fibrinolysis. 6 TAFIa operates by continuously removing C-terminal lysine residues on plasmin-modified partially degraded fibrin, thus attenuating the rate of plasminogen activation and fibrinolysis.…”