Vertex-Specific Proteins pUL17 and pUL25 Mechanically Reinforce Herpes Simplex Virus Capsids

Abstract

Using atomic force microscopy imaging and nanoindentation measurements, we investigated the effect of the minor capsid proteins pUL17 and pUL25 on the structural stability of icosahedral herpes simplex virus capsids. pUL17 and pUL25, which form the capsid vertex-specific component (CVSC), particularly contributed to capsid resilience along the 5-fold and 2-fold but not along the 3-fold icosahedral axes. Our detailed analyses, including quantitative mass spectrometry of the protein composition of the capsids, revealed that both pUL17 and pUL25 are required to stabilize the capsid shells at the vertices. This indicates that herpesviruses withstand the internal pressure that is generated during DNA genome packaging by locally reinforcing the mechanical sturdiness of the vertices, the most stressed part of the capsids.IMPORTANCE In this study, the structural, material properties of herpes simplex virus 1 were investigated. The capsid of herpes simplex virus is built up of a variety of proteins, and we scrutinized the influence of two of these proteins on the stability of the capsid. For this, we used a scanning force microscope that makes detailed, topographic images of the particles and that is able to perform mechanical deformation measurements. Using this approach, we revealed that both studied proteins play an essential role in viral stability. These new insights support us in forming a complete view on viral structure and furthermore could possibly help not only to develop specific antivirals but also to build protein shells with improved stability for drug delivery purposes.

Keywords: AFM; capsid; herpes simplex virus; stability.

FIG 1

Atomic force microscopy imaging of HSV-1 capsids. (a) Schematic of the HSV-1 capsid vertex region (41, 44). The UL25 part of the CVSC is proposed to be closest to the vertex, likely touching it (45). (b) AFM images of HSV-1 capsids. Based on the facet orientation and capsomer morphology, particles deposited on the 2-, 3-, and 5-fold icosahedral symmetry axes can be distinguished. Bar, 50 nm.

FIG 2

Both the CVSCs pUL17 and pUL25 contribute to the mechanical vertex stabilization of HSV-1 capsids. (a) Frequency distributions of particle spring constants (k) for particles with or without the CVSC, showing the shift to lower k values for the latter particles. (b) The average spring constants (k) for each orientation are shown for all three capsid types, comparing capsids in UL17- or UL25-null backgrounds to WT capsids. Error bars represent standard errors of the means, and the numbers of particles per type/orientation are indicated in white on each bar.

FIG 3

Protein copy numbers on capsids. Shown are quantitative mass spectrometry results for the abundance of pUL38, pUL18, pUL25, and pUL17 on the different capsids. The relative numbers of PSMs (67) are indicated on the y axis.