I also appreciate the guidance of my advisors, Professors David Dawson and David Chan, in the UCLA-Caltech Medical Scientist Training Program. Furthermore, I appreciate the support of the NIH NIAID F30 Fellowship, Josephine de Karman Fellowship, and Chen Graduate Innovator Award to pursue my research ideas. Finally, I thank my parents, Denise and Robert, for their dedication to helping others and for inspiring me to pursue a career in science and medicine to make a positive impact on the world.
We demonstrated that affinity maturation of the light chain variable domain is important for strong binding of VH3-23/VK1-5 neutralizing antibodies to Zika virus envelope domain III (EDIII) and identified interactions that contribute to weak, cross-reactive binding to West Nile Virus EDIII. Coronaviruses are another group of viruses responsible for widespread morbidity and mortality, including the severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East Respiratory Syndrome coronavirus (MERS-CoV) epidemics and current SARS-CoV-2 pandemic. Given concerns about new SARS-CoV-2 variants and the possibility for additional zoonotic betacoronaviruses to cause future outbreaks, we investigated how the epitopes on the SARS-CoV-2 receptor binding domain (RBD) represented by VH3-30- derived antibodies are targeted correlates with their neutralization potency and breadth of betacoronavirus recognition.
How antibodies recognize pathogenic viruses: structural correlates of antibody neutralization of HIV-1, SARS-CoV-2, and Zika. VH3-30 ANTIBODIES TARGETING THE SARS-COV-2RBD SHOW DIVERSE NEUTRALIZATION AND CROSS-REACTIVITY PROFILES.
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Abs targeting the SARS-CoV-2 RBD are an important component of the protective immune response against infection. We showed that these Abs bound varied epitopes on the SARS-CoV-2 spike RBD and could cause dissociation of the spike trimer. We show that affinity maturation of the light chain (LC) variable domain (VL) is important for strong binding of the repeating VH3-23/VK1-5 neutralizing Abs to ZIKV EDIII and identify interacting residues that contribute to weak, cross-reactive binding to the West -Nile virus.
Here we report binding and structural studies to gain insight into the affinity maturation and cross-reactivity of the VH3-23/VK1-5 class of anti-ZIKV Abs. There are 16 amino acid differences in the VH and nine differences in the VL of the Z004mature and Z004iGL. Mature IgGs showed weak binding to some of the EDIIIs; specifically Z004mature, Z006mature and Z034mature with DENV2 EDIII, Z004mature.
Perhaps correlating with the low affinity of the complex, the crystallographic asymmetric unit contained one Fab–WNV EDIII complex and four unbound Fabs (Figure 2.6A; SI Appendix , Table S2.1 ). By comparing binding of mature and iGL VH3-23/VK1-5 Ab to EDIIIs, we identified mutations introduced by somatic hypermutation that facilitate tight binding of Z004 Ab to ZIKV. A better understanding of the differences in interactions that contribute to neutralization versus cross-reactivity that leads to ADEs may enable strategic immunogen design.
This gave a partial solution with two Z004mature Fab–DENV1 EDIII molecules in the asymmetric unit, with one of the Fabs containing a misaligned constant (CHCL) domain. EDIII residues within 4 Å of VHVL are shown in magenta with the EK motif in blue. Sera from six immunization groups were tested; the key is shown in the legend at the top of the figure.
The Z004mature Fab–DENV1 EDIII (green–orange) from the crystal structure is superimposed on the EDIIIs of the mature ZIKV cryo-EM structure (PDB 6CO8) corresponding to the two-, three-, and fivefold local symmetry environments (white) . Single-particle cryo-electron microscopy structures of Fab-spike complexes showed that Fabs from five weakly neutralizing and one strongly neutralizing VH3-30 antibody caused dissociation from the peak trimer, with the strongly neutralizing antibody binding to a class 1 hybrid. 4 epitope. Surface views are shown for the side of the RBD facing the trimer center.
The weakly neutralizing Ab C133, which binds the lowest class 4 epitope in the RBD, showed the most extensive cross-reactivity with binding to all RBDs, although binding to SARS-CoV and Rs4081 was relatively reduced. The only two amino acids that differ between SARS-CoV and WIV1 that are close to the RBD-binding interface of the six VH3-30 Abs we characterized are S373RBD (a residue near the interface) and T430RBD (an interacting residue) (numbering based on SARS-CoV-2 RBD) ( SI Appendix , Figure S5.3 , S. The only two amino acids that differ between SARS-CoV and WIV1 that are close to the RBD-binding interface of the six VH3-30 Abs we characterized are S373RBD (residues near the interface ) and T430RBD (a interacting residues) (count based on SARS-CoV-2 RBD) ( SI Appendix , Figure S5.2 , S .
While bacteriophage P1 controls could be detected with the FITC filter, the signal was too low to detect VLPs in the sediment sample; .. the signal in the FITC filter using BONCAT in solution was even lower than when using anode disks.
BIBLIOGRAPHY
Structures of the Zika virus envelope protein and its complex with a flavivirus broadly protective antibody. Development of a potent and protective germline-like antibody against Zika virus in a recovered human. Immunization of Zika virus envelope protein domain III induces specific and neutralizing immune responses against Zika virus.
Virus-like particles displaying the envelope protein domain of Zika virus III induce potent neutralizing immune responses in mice. Prior dengue infection is associated with increased viral load in patients infected with dengue but not Zika. A protective E-dimer-based Zika virus subunit vaccine designed to reverse antibody-dependent enhancement of dengue infection.
Pichia pastoris-expressed Zika virus envelope domain III on a virus-like particle platform: design, production, and immunological evaluation. ENCLOSURED VIRUS-LIKE PARTICLES (eVLPs) EXPRESSING MODIFIED FORMS OF ZIKA VIRUS PROTEINS E AND NS1 PROTECT MICE FROM ZIKA VIRUS INFECTION.
