An anti-TfR1 Fab fragment that blocks the cell entry of a family of New World Hemorrhagic Fever Arenaviruses: Functional evaluation, structural analysis and antibody-antigen docking modeling

PENICHET M. L.; SAWAYA, MICHAEL R.; DIEP, A.; RODRÍGUEZ J. A.; CASCIO, DUILIO; ZIEGENBEIN, J.; HELGUERA, G.; DANIELS-WELLS T.; ARAGÜES, R.; PAYÉS, C.

San Diego

Encuentro; Annual Meeting of the Antibody Society: Antibody Engineering and Therapeutics; 2017

The Antibody Society

There arefive New World Hemorrhagic Fever (NWHF) that cause grave human diseases withhigh fatality rate and limited therapeutic options. Previously, we observedthat the recombinant antibody ch128.1 specific for hTfR1 can block the entry ofall NWHF arenaviruses that use this receptor as a gateway of entry: Machupo(MACV), Chapare (CHAV), Guanarito (GTOV), Sabiá (SABV) and Junín (JUNV). Thesurface glycoprotein (GP) of all pathogenic NWHF arenaviruses mediate humancell binding through the apical domain of hTfR1. To better understand thestructural basis of the antibody-antigen interaction and the blockade of the GPArenavirus binding to TfR1 we performed functional and X-ray structuralanalysis of Fab fragment of ch128.1 IgG1 (Fab128.1), followed by computationaldocking analysis of Fab128.1 and hTfR1. Here we report that the Fab fragment ofch128.1 IgG1 (Fab128.1) is sufficient to significantly block theinternalization of pseudoviruses decorated with GP1/GP2 of the pathogenicarenaviruses MACV, CHAV, GTOV, SABV and JUNV. Structural analysis of Fab128.1crystals was performed with X-ray diffraction collected from a synchrotronbeam, and the atomic structure solved by molecular replacement with 2.69Åresolution of the complementary determinant regions (CDRs). With the Fab128.1structural data and the hTfR1 crystal structure complexed to MACV-GP1 (pdb3KAS) we performed a computational docking analysis to obtain a structuralmodel of the Fab-antigen complex. We started with a rigid-body direct method tofind the structure of the target complex located at the minimum of Gibbs freeenergy in the conformational space using the ClusPro package. We included forthe analysis an attraction mask of surface residues based on our TfR1mutagenesis analysis (S338-S368) that identified residues critical for ch128.1arenaviral entry blockade, obtaining clusters of Fab-antigen complexes. Thefour largest cluster models where used as putative antibody-antigen input for aflexible-backbone docking analysis with Rosetta SnugDock. The antibody-antigenmodel with the lowest interface score obtained from the largest ClusPro clustermatched the mutagenesis data and predicted binding to at least 25 TfR1residues. Moreover, since at least 10 residues on the surface of TfR1 thatinteracted with Fab128.1 overlap with the MACV-GP1 binding site, this model mayexplain the blockade of the pathogenic arenavirus internalization. Furthermutagenesis studies on the variable region of Fab128.1 are required to confirmthe structural predictions.  Thesestudies suggest that modeling using computational docking analysis haspotential predictive power to understand the blocking functional activity ofFab128.1, which may lead to a rational design of a safer and more effectivebiotherapeutic against NWHFs.