The TFPIα C-terminal tail is essential for TFPIα-FV-short-protein S complex formation and synergistic enhancement of TFPIα.
J Thromb Haemost
; 21(12): 3568-3580, 2023 12.
Article
in En
| MEDLINE
| ID: mdl-37739040
ABSTRACT
BACKGROUND:
For maximal TFPIα functionality, 2 synergistic cofactors, protein S and FV-short, are required. Both interact with TFPIα, protein S through Kunitz 3 residues Arg199/Glu226 and FV-short with the C-terminus. How these interactions impact the synergistic enhancement remains unclear.OBJECTIVES:
To determine the importance of the TFPIα-protein S and TFPIα-FV-short interactions for TFPIα enhancement.METHODS:
TFPIα variants unable to bind protein S (K3m [R199Q/E226Q]) or FV-short (ΔCT [aa 1-249]) were generated. TFPIα-FV-short binding was studied by plate-binding and co-immunoprecipitation assays; functional TFPIα enhancement by FXa inhibition and prothrombin activation.RESULTS:
While WT TFPIα and TFPIα K3m bound FV-short with high affinity (Kdâ¼2nM), TFPIα ΔCT did not. K3m, in contrast to WT, did not incorporate protein S in a TFPIα-FV-short-protein S complex while TFPIα ΔCT bound neither FV-short nor protein S. Protein S enhanced WT TFPIα-mediated FXa inhibition, but not K3m, in the absence of FV-short. However, once FV-short was present, protein S efficiently enhanced TFPIα K3m (EC50 4.7nM vs 2.0nM for WT). FXa inhibition by ΔCT was not enhanced by protein S alone or combined with FV-short. In FXa-catalyzed prothrombin activation assays, FV-short enhanced TFPIα K3m function in the presence of protein S (5.5 vs 10.4-fold enhancement of WT) whereas ΔCT showed reduced or lack of enhancement by FV-short and protein S, respectively.CONCLUSION:
Full TFPIα function requires the presence of both cofactors. While synergistic enhancement can be achieved in the absence of TFPIα-protein S interaction, only TFPIα with an intact C-terminus can be synergistically enhanced by protein S and FV-short.Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Blood Coagulation
/
Prothrombin
Limits:
Humans
Language:
En
Journal:
J Thromb Haemost
Journal subject:
HEMATOLOGIA
Year:
2023
Document type:
Article
Affiliation country:
Reino Unido