5nmv
Crystal structure of 2F22 Fab fragment against TFPI1
Structural highlights
FunctionTFPI1_HUMAN Inhibits factor X (X(a)) directly and, in a Xa-dependent way, inhibits VIIa/tissue factor activity, presumably by forming a quaternary Xa/LACI/VIIa/TF complex. It possesses an antithrombotic action and also the ability to associate with lipoproteins in plasma. Publication Abstract from PubMedBACKGROUND: Initiation of coagulation is induced by binding of activated factor VII (FVIIa) to tissue factor (TF) and activation of factor X (FX) in a process regulated by tissue factor pathway inhibitor (TFPI). TFPI contains three Kunitz-type protease inhibitor domains (K1-K3) of which K1 and K2 block the active sites of FVIIa and FXa respectively. OBJECTIVE: To produce a monoclonal antibody (mAb) directed towards K1, to characterize the binding epitope, and to study its effect on TFPI inhibition. METHODS: A monoclonal antibody, mAb2F22, was raised against the N-terminal TFPI(1-79) fragment. Binding data were obtained by surface plasmon resonance analysis. The Fab-fragment of mAb2F22, Fab2F22, was expressed and the structure of its complex with TFPI(1-79) determined by X-ray crystallography. Effects of mAb2F22 on TFPI inhibition were measured in buffer- and plasma- based systems. RESULTS: mAb2F22 bound exclusively to K1 of TFPI (KD ~1 nM) and not to K2. The crystal structure of Fab2F22/TFPI (1-79) mapped an epitope on K1 including seven residues upstream of the domain. TFPI inhibition of TF/FVIIa amidolytic activity was neutralized by mAb2F22, although the binding epitope on K1 did not include the P1 residue. Binding of mAb2F22 to K1 blocked TFPI inhibition of the FXa amidolytic activity and normalised haemostasis in haemophilia human A-like plasma and whole blood. CONCLUSION: mAb2F22 blocked TFPI inhibition of both FVIIa and FXa activities and mapped a FXa exosite for binding to K1. It reversed TFPI feed-back inhibition of TF/FVIIa-induced coagulation and restored clot formation in FVIII-neutralized human plasma and blood. This article is protected by copyright. All rights reserved. Factor Xa and VIIa inhibition by Tissue Factor Pathway Inhibitor is prevented by a monoclonal antibody to its Kunitz-1 domain.,Augustsson C, Svensson A, Kjaer B, Chao TY, Wenjuan X, Krogh BO, Breinholt J, Clausen JT, Hilden I, Petersen HH, Petersen LC J Thromb Haemost. 2018 Mar 12. doi: 10.1111/jth.14000. PMID:29532595[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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