Selective modulation of thrombin‐activatable fibrinolysis inhibitor (TAFI) activation by thrombin or the thrombin‐thrombomodulin complex using TAFI‐derived peptides

• Published in: Journal of thrombosis and haemostasis Vol. 13; no. 11; pp. 2093 - 2101
• Main Authors: Plug, T., Marquart, J. A., Marx, P. F., Meijers, J. C. M.
• Format: Journal Article
• Language: English
• Published: England Elsevier Limited 01.11.2015
• Subjects: Amino Acid Sequence; carboxypeptidase B2; Carboxypeptidase B2 - antagonists & inhibitors; Carboxypeptidase B2 - chemistry; Enzyme Activation - drug effects; fibrinolysis; Half-Life; Humans; Inhibitory Concentration 50; Models, Molecular; Molecular Sequence Data; Peptide Fragments - chemical synthesis; Peptide Fragments - pharmacology; Protein Binding; Protein Conformation; Structure-Activity Relationship; Surface Plasmon Resonance; thrombin; Thrombin - metabolism; Thrombin - pharmacology; thrombin‐activatable fibrinolysis inhibitor; thrombomodulin; Thrombomodulin - metabolism; fibrinolysis; thrombin-activatable fibrinolysis inhibitor; thrombin; carboxypeptidase B2; thrombomodulin
• DOI: 10.1111/jth.13133
• ISSN: 1538-7933; 1538-7836

Summary Background Thrombin‐activatable fibrinolysis inhibitor (TAFI) is a risk factor for coronary heart disease. TAFI is proteolytically activated by thrombin, the thrombin‐thrombomodulin complex and plasmin. Once active, it dampens fibrinolysis and inflammation. The aim of this study was to generate TAFI‐derived peptides that specifically modulate TAFI activation and activity. Methods Thirty‐four overlapping TAFI peptides, and modifications thereof, were synthesized. The effects of these peptides on TAFI activation and TAFIa activity were determined. In addition, the binding of the peptides to thrombin were determined. Results Four peptides (peptides 2, 18, 19 and 34) inhibited TAFI activation and two peptides (peptides 14 and 24) inhibited TAFIa activity directly. Peptide 2 (Arg12‐Glu28) and peptide 34 (Cys383‐Val401) inhibited TAFI activation by the thrombin‐thrombomodulin complex with IC50 values of 7.3 ± 1.8 and 6.1 ± 0.9 μm, respectively. However, no inhibition was observed in the absence of thrombomodulin. This suggests that the regions Arg12‐Glu28 and Cys383‐Val401 in TAFI are involved in thrombomodulin‐mediated TAFI activation. Peptide 18 (Gly205‐Ser221) and peptide 19 (Arg214‐Asp232) inhibited TAFI activation by thrombin and the thrombin‐thrombomodulin complex. Furthermore, these peptides bound to thrombin (KD: 1.5 ± 0.4 and 0.52 ± 0.07 μm for peptides 18 and 19, respectively), suggesting that Gly205‐Asp232 of TAFI is involved in binding to thrombin. Peptide 14 (His159‐His175) inhibited TAFIa activity. The inhibition was TAFIa specific, because no effect on the homologous enzyme carboxypeptidase B was observed. Conclusions Thrombin‐activatable fibrinolysis inhibitor‐derived peptides show promise as new tools to modulate TAFI activation and TAFIa activity. Furthermore, these peptides revealed potential binding sites on TAFI for thrombin and the thrombin‐thrombomodulin complex.