Open AccessCrystal structure, epitope, and functional impact of an antibody against a superactive FVII a provide insights into allosteric mechanism
Author(s) -
Jiang Longguang,
Xie Xie,
Li Jinyu,
Persson Egon,
Huang Mingdong
Publication year - 2019
Publication title -
research and practice in thrombosis and haemostasis
Language(s) - English
Resource type - Journals
ISSN - 2475-0379
DOI - 10.1002/rth2.12211
Subject(s) - epitope , allosteric regulation , chemistry , epitope mapping , surface plasmon resonance , docking (animal) , binding site , rational design , computational biology , biophysics , protein structure , recombinant dna , antigen , biochemistry , receptor , nanotechnology , biology , medicine , genetics , gene , materials science , nursing , nanoparticle
Background Blood coagulation factor VII a ( FVII a) plays its critical physiological role in the initiation of hemostasis. Even so, recombinant FVII a is successfully used as a bypassing agent for factor VIII or IX in the treatment of bleeds in patients with severe hemophilia with inhibitors. To investigate the utility of more potent FVII a variants with enhanced intrinsic activity, molecules such as V21D/E154V/M156Q‐ FVII a ( FVII a DVQ ) were designed. Methods Surface plasmon resonance was used to characterize the binding of mA b4F5 to FVII a DVQ and related variants. X‐ray crystallography was used to determine the structure of the Fab fragment of mA b4F5 (Fab4F5). Molecular docking and small angle X‐ray scattering led to a model of FVII a DVQ :Fab4F5 complex. Results The binding experiments, functional effects on FVII a DVQ and structure of mA b4F5 (originally intended for quantification of FVII a DVQ in samples containing FVII (a)) pinpointed the epitope (crucial role for residue Asp21) and shed light on the role of the N‐terminus of the protease domain in FVII a allostery. The potential antigen‐combining sites are composed of 1 hydrophobic and 1 negatively charged pocket formed by 6 complementarity‐determining region (CDR) loops. Structural analysis of Fab4F5 shows that the epitope interacts with the periphery of the hydrophobic pocket and provides insights into the molecular basis of mA b4F5 recognition and tight binding of FVII a DVQ . Conclusion The binary complex explains and supports the selectivity and functional consequences of Fab4F5 association with FVII a DVQ and illustrates the potentially unique antigenicity of this FVII a variant. This will be useful in the design of less immunogenic variants.