The structure of HSV-1 gB in complex with D48 Fab and structural and sequence comparison with HCMV and EBV gB with their neutralization antibody reveal DII as a conservative antigenic domain across herpesviruses. (IMAGE)
Caption
(A) Cryo-EM structure overview of HSV-1 gB in complex with D48 Fab. The side view and top view were presented. The HSV-1 gB trimer and D48 Fab VH/VL were colored differently.
(B) Neutralization assay of D48 Fab, D48 IgG1 and control IgG to HSV-1 infection of Vero cells. The neutralization (%) was calculated by [(mean plaque number of blank neutralization - plaque number of certain antibody neutralization at certain concentration) / mean plaque number of blank neutralization] * 100(%). Half maximal inhibitory concentrations (IC50) were calculated by non-linear regression for each antibody. Data points were shown as the mean ± SEM from three replicate wells.
(C) Biolayer interferometry (BLI) kinetic assay to D48 Fab binding to HSV-1 gB. Different concentrations were marked beside the corresponding curve in different colors. Kinetic data from one experiment was shown.
(D) Sequence and domain segmentation of HSV-1 gB. Domains were highlighted in different colors and interval residue positions were marked.
(E) Ribbon structure for a monomer of HSV-1 gB in complex with D48 Fab. The gB domain colors were consistent with (D).
(F) Zoomed-in view shows polar interaction diagram for HSV-1 gB DII domain with D48 Fab VL. The interaction residues were shown as sticks. The three CDRs and FR2 were colored differently and zoom views for LCDR1, LCDR3, and FR2 were outlined.
(G) Details for HSV-1 gB polar interaction residues with D48 Fab and the residue-mutant gB proteins affinity with D48 Fab measured by BLI. The listed polar interaction included hydrogen bonds (H-bonds) and salt bridges. The affinity was represented by the KD reported by BLI assays on wildtype (WT) or alanine-mutant HSV-1 gB binding to D48 Fab. Deeper red meant stronger affinity and deeper blue meant weaker.
(H) Detailed interaction diagram for HSV-1 gB polar interaction with D48 Fab in separate zoom view for LCDR3, LCDR1, and FR2. Key interacting residues of D48 or HSV-1 gB were marked in color consistently to (F).
(I) Comparison of architecture and antibody binding footprint of HSV-1, HCMV, and EBV post-fusion gB in complex with its DII-specific antibody. The VH and VL of D48, SM5-1, and 3A3 were colored differently. The antibody footprints on the DII domain were consistently colored by VH or VL.
(J) Sequence alignment of eight different gB DII domains across α, β, and γ herpesviruses. The secondary structure annotation was generated in the template of HSV-1 gB. Identical residues are shown as white symbols on red background, and similar residues are shown as red symbols. The exact residues of antibody footprint on DII domains were marked as arrows and colored by the corresponding binding antibody VH or VL in (I).
(K) Comparison of the architecture of HSV-1, HCMV, and EBV pre-fusion gB in complex with its DII-specific antibody generated by pre-post DII domain alignment. The VH and VL of D48, SM5-1, and 3A3 were colored differently as (I). The domain color principle for cartoon diagrams for pre-fusion gB in complex with antibody was the same as (E).
Credit
Cong Sun, Sun Yat-sen University Cancer Center
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