Structural characterization of a neutralizing mAb H16.001, a potent candidate for a common potency assay for various HPV16 VLPs

Abstract

With more human papillomavirus (HPV) virus-like particle (VLP) vaccines to hit the market in future, a monoclonal antibody (mAb) with preferably comparable reactivity against vaccines from different expression systems and bioprocesses is urgently needed for the potency characterization. Among all mAbs against HPV16 collected, rabbit mAb H16.001 is potently neutralizing with the highest affinity, recognizes an immune-dominant epitope, and can comparably react with HPV16 vaccines from various sources. Cryo-electron microscopic (cryo-EM) structure demonstrated that 360 H16.001 Fabs could bind to HPV16 capsid in preferable binding manner without steric hindrance between neighboring Fabs, potentially supporting its identification for VLP structural integrity and utility in monitoring VLP structural probity. This structural analysis indicated that mAb H16.001 afforded unbiased potency characterization for various HPV16 vaccines and was potential for use in vaccine regulation practice. This study also showed a model process for selecting suitable mAbs for potency assays of other vaccines.

Cryo EM Analysis Reveals Inherent Flexibility of Authentic Murine Papillomavirus Capsids

“…Previously, reconstructions of papillomaviruses with full icosahedral symmetry imposed have produced maps limited to modest resolution; however, recently subparticle reconstruction approaches have been successfully used to obtain high resolution [ 16 , 17 , 36 ]. Consistent with this finding, we found that for the native papillomavirus the icosahedrally enforced reconstruction did not advance beyond 4.4 Å resolution.…”

B-cell epitope discovery: The first protein flexibility-based algorithm–Zika virus conserved epitope demonstration

“…To better understand the generality of flexibility-based method presented here, protein flexibility's influence on conserved human papillomavirus [16] (HPV) epitopes was examined. Benchmarking was performed against two highly conserved (mean residue evolutionary rate of 1.52) [25] HPV epitopes (PDB IDs: 7CN2 [54], 3J8Z [55]) out of eight structurally characterized HPV epitopes (evolutionary rate range of 1.18-2.47). Like flaviviruses, linear combination of HPV isolated and VLP protein flexibility (using weights which minimized the root-meansquare difference between the two flexibility profiles) performed with the strongest capacity (of all metrics examined) to pinpoint conserved HPV epitope residues (a PRAUC mean 0.21 above the value of no discrimination, 0.07) (Table 3).…”

“…Of the seven highly conserved epitopes, 14% were associated with ZIKV, 0% were associated with dengue virus serotype 1, 14% were associated with dengue virus serotype 2, 14% were associated with dengue virus serotype 3, 29% were associated with dengue virus serotype 4, ROCAUC and PRAUC values calculated against two highly conserved HPV epitopes [54,55] are shown for the linear difference of isolated and VLP RMSF (fmon-fvlp),…”

“…HPV epitopes were aligned to the sequence of L1 protein chain A (PDB ID: 5KEP). Two structurally characterized epitopes with low mean residue evolutionary rates (PDB IDs: 7CN2 [54], 3J8Z [55]) out of eight epitopes were compiled for benchmarking.…”

“…An analogous process was used for HPV epitopes. Positives were defined as residues from two highly conserved HPV epitopes (PDB IDs: 7CN2 [54], 3J8Z [55]) out of eight epitopes. Negatives were defined as all other HPV residues in L1 protein chain A (PDB ID: 5KEP) (Table 3).…”