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• W2022469057 abstract "Background Down-regulation of fibrinolysis due to cleavage of C-terminal lysine residues from partially degraded fibrin is mainly exerted by the carboxypeptidase activity of activated thrombin-activatable fibrinolysis inhibitor (TAFIa). Recently, some intrinsic carboxypeptidase activity (i.e. zymogen activity) was reported for the proenzyme (TAFI); however, there is some discussion about its ability to cleave high molecular weight substrates. Objective We aimed to identify and characterize nanobodies toward mouse TAFI (mTAFI) that stimulate the zymogen activity and to test their effect in an in vitro clot lysis assay and an in vivo mouse thromboembolism model. Methods and results Screening of a library of nanobodies toward mTAFI revealed one nanobody (VHH-mTAFI-i49) that significantly stimulates the zymogen activity of mTAFI from undetectable (< 0.35 U mg−1) to 4.4 U mg−1 (at a 16-fold molar ratio over mTAFI). The generated carboxypeptidase activity is unstable at 37 °C. Incubation of mTAFI with VHH-mTAFI-i49 revealed a time-dependent reduced activatability of mTAFI. Epitope mapping revealed that Arg227 and Lys212 are important for the nanobody/mTAFI interaction and suggest destabilization of mTAFI by disrupting the stabilizing interaction between the activation peptide and the dynamic flap region. In vitro clot lysis experiments revealed an enhanced clot lysis due to a reduced activation of mTAFI during clot formation. In vivo application of VHH-mTAFI-i49 in a mouse thromboembolism model decreased dose-dependently the fibrin deposition in the lungs of thromboembolism-induced mice. Conclusion The novel, nanobody-induced, reduced activatability of mTAFI demonstrates to be a very potent approach to enhance clot lysis." @default.
• W2022469057 created "2016-06-24" @default.
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• W2022469057 date "2014-02-01" @default.
• W2022469057 modified "2023-09-27" @default.
• W2022469057 title "Identification of a novel, nanobody‐induced, mechanism of TAFI inactivation and its in vivo application" @default.
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• W2022469057 doi "https://doi.org/10.1111/jth.12473" @default.
• W2022469057 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/24354544" @default.
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