Cryo-EM structure of a herpesvirus capsid at 3.1 Å

INTRODUCTION Herpes simplex virus type 2 (HSV-2) is a sexually transmitted virus and is the leading causative agent of genital ulcer disease (GUD) worldwide. Patients with HSV-2 have a higher risk of acquiring human immunodeficiency virus (HIV) infection. HSV-2, as well as the closely related herpes simplex virus 1 (HSV-1), are simplexviruses with a natural-host range restricted to humans, belonging to the family of Herpesviridae , whose other members are responsible for a number of diseases, including congenital disorders (e.g., human cytomegalovirus) and even cancers (e.g., Epstein-Barr virus and Kaposi sarcoma herpesvirus). HSVs’ ability to establish a lifelong latent infection within hosts and recurrent reactivation from latency make them highly effective pathogens with seropositivity rates close to 100% in adult populations. RATIONALE The herpesvirus virion is genetically and structurally one of the largest and most complex viruses known. It has a T = 16 (triangulation number) icosahedral capsid with a diameter of ~125 nm that not only protects the viral genome physically from damage but also plays an important role in the release of viral genome into the nucleus of the host cell. HSV capsid assembly requires the ordered packing of about 4000 protein subunits into the hexons, pentons, and triplexes that comprise the capsid. Previous studies have suggested that the directionality of triplexes on the capsid shell and disulfide bond formation between capsid proteins contribute to HSV capsid assembly, but in the absence of an atomic description of HSV capsids, the molecular basis that drives capsid assembly has remained elusive. RESULTS By using a “block-based” image reconstruction approach combined with a Ewald sphere correction, we have visualized the HSV capsid at 3.1-A resolution by cryo–electron microscopy (cryo-EM) and have built an atomic structure, which includes 28,138 residues in the asymmetric unit, belonging to 46 different conformers of four capsid proteins (VP5, VP23, VP19C, and VP26). These organize into three types of hexons (central, peripentonal, and edge) that contain the major capsid protein VP5 and the small capsid protein VP26, pentons made up of VP5, and triplexes composed of VP23 and VP19C. Acting as core organizers, VP5 proteins form extensive intermolecular networks, involving disulfide bonds (25 per asymmetric unit) and noncovalent interactions, with VP26 proteins and triplexes, that underpin capsid stability and assembly. Together with previous low-resolution structural results, we propose a model for the ordered assembly of the capsid using basic assembly units (a triplex and its covalently linked lasso triangle formed by three VP5s), which then cluster into higher-order structures conforming to twofold symmetry and guide nascent assembly intermediates into the correct T = 16 geometry. CONCLUSION The marked improvement in the resolution of the structure of the herpesvirus capsid determined by cryo-EM allows the first steps toward understanding the drivers of assembly and the basis of stability of the capsid. In addition, the atomic structure could guide rational design of therapeutic agents for treating tumors and therapeutic strategies against HSV.