Heterogeneity of HPV16 virus-like particles indicates a complex assembly energy surface

• Published in: Virology (New York, N.Y.) N.Y.), 2024-12, Vol.600, p.110211, Article 110211
• Main Authors: Patterson, Angela, Young, Kim, Biever, MacRyan P., Klein, Shelby M., Huang, Sheng-Yuan, DePhillips, Pete A., Jacobson, Stephen C., Jarrold, Martin F., Zlotnick, Adam
• Format: Article
• Language: English
• Subjects: Capsid; Capsid - chemistry; Capsid - metabolism; Capsid Proteins - chemistry; Capsid Proteins - genetics; Capsid Proteins - metabolism; Charge detection-mass spectrometry; HPV16; Human papilloma virus; Human papillomavirus 16 - chemistry; Human papillomavirus 16 - genetics; Humans; Hydrogen-Ion Concentration; Oncogene Proteins, Viral - chemistry; Oncogene Proteins, Viral - genetics; Oncogene Proteins, Viral - metabolism; Osmolar Concentration; Oxidation-Reduction; Resistive pulse sensing; Self-assembly; Virion - chemistry; Virion - metabolism; Virus Assembly
• ISSN: 0042-6822; 1096-0341; 1096-0341
• DOI: 10.1016/j.virol.2024.110211

Human Papillomavirus serotype 16 (HPV16) capsid protein (L1) pentamers canonically assemble into T = 7 icosahedral capsids. Such virus-like particles are the basis of the HPV vaccine. We examined assembly of L1 pentamers in response to pH, mild oxidants, and ionic strength and found a mixture of closed, roughly spherical structures from ∼20 to ∼70 nm in diameter, indicating the presence of many kinetically accessible energy minima. Using bulk and single particle techniques we observed that the size distribution changes but does not reach homogeneity. Though heterogenous in size, particles showed uniform responses to low ionic strength dissociation, thermal unfolding, and susceptibility to protease digestion. These assays suggest maturation over time, but at different rates. Cysteine oxidation further stabilized particles at early, but not late, times without changing general characteristics including thermal stability and protease digestion. These data show complex assembly paths to species of different sizes, but with locally similar interactions. [Display omitted] •Assembly of HPV16 L1 pentamers results in a heterogeneous mixture of virus-like particles.•Single particle methods allow quantification of the distribution of species and how this distribution changes over time.•Though particles have different sizes, their biophysical properties are similar. This implies similar local structure.