Maturation of the Human Papillomavirus 16 Capsid

Abstract

Papillomaviruses are a family of nonenveloped DNA viruses that infect the skin or mucosa of their vertebrate hosts. The viral life cycle is closely tied to the differentiation of infected keratinocytes. Papillomavirus virions are released into the environment through a process known as desquamation, in which keratinocytes lose structural integrity prior to being shed from the surface of the skin. During this process, virions are exposed to an increasingly oxidative environment, leading to their stabilization through the formation of disulfide cross-links between neighboring molecules of the major capsid protein, L1. We used time-lapse cryo-electron microscopy and image analysis to study the maturation of HPV16 capsids assembled in mammalian cells and exposed to an oxidizing environment after cell lysis. Initially, the virion is a loosely connected procapsid that, under in vitro conditions, condenses over several hours into the more familiar 60-nm-diameter papillomavirus capsid. In this process, the procapsid shrinks by ~5% in diameter, its pentameric capsomers change in structure (most markedly in the axial region), and the interaction surfaces between adjacent capsomers are consolidated. A C175S mutant that cannot achieve normal inter-L1 disulfide cross-links shows maturation-related shrinkage but does not achieve the fully condensed 60-nm form. Pseudoatomic modeling based on a 9-Å resolution reconstruction of fully mature capsids revealed C-terminal disulfide-stabilized “suspended bridges” that form intercapsomeric cross-links. The data suggest a model in which procapsids exist in a range of dynamic intermediates that can be locked into increasingly mature configurations by disulfide cross-linking, possibly through a Brownian ratchet mechanism.

The U4 Antibody Epitope on Human Papillomavirus 16 Identified by Cryo-electron Microscopy

“…Subsequent structural studies have refined the C-terminal arm arrangement in HPV capsids (Fig. 8A) (9,17,18). Of the two types of available binding spaces (defined as type I and II in Results) for H16.U4 (Fig.…”

“…This combination of factors resulted in the lack of full Fab density in the complex map after the reconstruction process. (18) that expose the H16.U4 epitope were defined as area I, bordered by three hexavalent capsomers (black hexagons), and area II that is surrounded by two hexavalent capsomers (black hexagons) and one pentavalent capsomer (black pentagon). The correct orientation along the pseudodyad axis for the fitted Fab was identified according to steric interference between fitted molecules (Table 2).…”

“…The C terminus of each L1 protein, called the C-terminal arm, extends along the capsid floor to interact with the neighboring capsomer before returning to the original donor cap-somer (9,17,18). Intercapsomer disulfide bonds are formed between cysteine C428 and C175, which stabilize the icosahedral structure and play an important role in virus maturation (18,19). The core of the capsomer is composed of the common viral structural motif, the antiparallel ␤-strands BIDG and CHEF (20).…”

“…Twelve capsomers lie on an icosahedral five-fold vertex and are referred to as pentavalent capsomers, whereas the remaining 60 capsomers are each surrounded by six other capsomers and referred to as hexavalent capsomers. The C terminus of each L1 protein, called the C-terminal arm, extends along the capsid floor to interact with the neighboring capsomer before returning to the original donor cap-somer (9,17,18). Intercapsomer disulfide bonds are formed between cysteine C428 and C175, which stabilize the icosahedral structure and play an important role in virus maturation (18,19).…”

Kallikrein-8 Proteolytically Processes Human Papillomaviruses in the Extracellular Space To Facilitate Entry into Host Cells

“…In addition, papillomaviruses undergo, like many other viruses, a maturation process after initial assembly. During this maturation intermolecular disulfide bonds between L1 molecules are formed that covalently link adjacent pentameric capsomers (7,9,10).…”