Volume 19, Issue 5, Pages 696-704 (May 2016) Structures of the Zika Virus Envelope Protein and Its Complex with a Flavivirus Broadly Protective Antibody  Lianpan Dai, Jian Song, Xishan Lu, Yong-Qiang Deng, Abednego Moki Musyoki, Huijun Cheng, Yanfang Zhang, Yuan Yuan, Hao Song, Joel Haywood, Haixia Xiao, Jinghua Yan, Yi Shi, Cheng-Feng Qin, Jianxun Qi, George F. Gao  Cell Host & Microbe  Volume 19, Issue 5, Pages 696-704 (May 2016) DOI: 10.1016/j.chom.2016.04.013 Copyright © 2016 Elsevier Inc. Terms and Conditions

Cell Host & Microbe 2016 19, 696-704DOI: (10.1016/j.chom.2016.04.013) Copyright © 2016 Elsevier Inc. Terms and Conditions

Figure 1 Overall Structure of the ZIKV-E Protein (A) Schematic diagram of domain organization for ZIKV-E. Domain I (red), domain II (yellow), and domain III (blue). A 48-residue stem segment links the stably folded ZIKV-E ectodomain with the C-terminal transmembrane anchor. (B) Dimer structure of ZIKV-E. ZIKV-E has three distinct domains: β-barrel-shaped domain I, finger-like domain II, and immunoglobulin-like domain III. Domain II is responsible for the dimerization of ZIKV-E. The fusion loop is buried by the domains I and II of the other ZIKV-E monomer. Secondary structural elements are labeled. See also Figures S1 and S2 and Table S1. Cell Host & Microbe 2016 19, 696-704DOI: (10.1016/j.chom.2016.04.013) Copyright © 2016 Elsevier Inc. Terms and Conditions

Figure 2 Binding of ZIKV-E to a Broadly Neutralizing Antibody 2A10G6 (A) Gel filtration profile of ZIKV-E and 2A10G6 Fab. The ZIKV-E and 2A10G6 Fab proteins elute as single monomer peaks in the gel filtration curves, respectively. The ZIKV-E/2A10G6 Fab complex displays a shifted complex peak with an extra 2A10G6 Fab peak. All the samples were assessed by SDS-PAGE. (B) BIAcore diagram of 2A10G6 Fab bound to ZIKV-E protein. The 2A10G6 Fab binds to the ZIKV-E protein with a high affinity and slow kinetics. The KD value was calculated by the BIAcore 3000 analysis software (BIAevaluation Version 4.1). Cell Host & Microbe 2016 19, 696-704DOI: (10.1016/j.chom.2016.04.013) Copyright © 2016 Elsevier Inc. Terms and Conditions

Figure 3 Neutralization Activity and Protection of 2A10G6 against ZIKV (A) Neutralization activity of 2A10G6 to ZIKV. ZIKV was mixed with 5-fold serial dilutions of 2A10G6. Then neutralization activity was evaluated by plaque reduction assays in duplicates using BHK21 cells. Data are represented as mean ± SEM. The figure represents two independent experiments. (B) Therapeutic activity of 2A10G6 against ZIKV in A129 mice. 5-week-old A129 mice were inoculated with 105 PFU of ZIKV. At 24 hr post virus infection, mice were passively transferred a single inoculation of 500 μg antibody 2A10G6, or PBS as control. The survival of the mice was monitored until 21 days post infection. The survival curves were constructed using data with five mice for each arm. Cell Host & Microbe 2016 19, 696-704DOI: (10.1016/j.chom.2016.04.013) Copyright © 2016 Elsevier Inc. Terms and Conditions

Figure 4 Complex Structure of 2A10G6 Fab Bound to ZIKV-E (A) Overall complex structure. The 2A10G6 Fab binds to the domain II tip of ZIKV-E monomer at a perpendicular angle with the fusion loop and bc loop involved in the interaction. (B and C) Cartoon representations of interacting residues in the ZIKV-E (B) and 2A10G6 Fab (C). The fusion loop of ZIKV-E deeply inserts into a hole formed by both the heavy chain and light chain of 2A10G6, while the bc loop contacts only the heavy chain. See also Figure S4. Cell Host & Microbe 2016 19, 696-704DOI: (10.1016/j.chom.2016.04.013) Copyright © 2016 Elsevier Inc. Terms and Conditions

Figure 5 Analysis of the Detailed Interaction between the 2A10G6 and ZIKV-E (A) Four polypeptide elements, three from the heavy chain (light pink) and one from the light chain (cyan), surround the domain II tip of ZIKV-E. (B) The heavy chain forms five hydrogen bonds (green dashed lines) with the ZIKV-E residues. The hydrophobic residues W101, L107, and F108 in the fusion loop contribute most of the contacts between 2A10G6 and ZIKV-E. Cell Host & Microbe 2016 19, 696-704DOI: (10.1016/j.chom.2016.04.013) Copyright © 2016 Elsevier Inc. Terms and Conditions

Figure 6 Comparison with Other Fusion Loop-Targeting Neutralizing Antibodies (A) Structure of prM-E heterodimer of dengue virus serotype 2 (DENV2) (PDB: 3C5X). The pr peptide is shown in green, with domain coloring described previously in Figure 1. (B) Structure of ZIKV-E/2A10G6 complex with coloring described previously in Figure 5. (C) Structure of fusion loop epitope (FLE)-specific E53 in complex with the West Nile virus E protein (PDB: 3I50). The heavy chain is in lemon, and the light chain is in wheat. (D) Structure of E-dimer epitope (EDE)-specific antibody A11 in complex with the DENV2 E protein (PDB: 4UT7). The heavy chain is in light gray, and the light chain is in light orange. See also Figures S2 and S3 and Table S2. Cell Host & Microbe 2016 19, 696-704DOI: (10.1016/j.chom.2016.04.013) Copyright © 2016 Elsevier Inc. Terms and Conditions