Biochemical and structural characterization of the capsid-bound tegument proteins of human cytomegalovirus

• Published in: Journal of structural biology Vol. 174; no. 3; pp. 451 - 460
• Main Authors: Yu, Xuekui, Shah, Sanket, Lee, Manfred, Dai, Wei, Lo, Pierrette, Britt, William, Zhu, Hua, Liu, Fenyong, Hong Zhou, Z.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.06.2011
• Subjects: Capsid - chemistry; Capsid - ultrastructure; Capsid Proteins - chemistry; Capsid Proteins - ultrastructure; Cells, Cultured; Chemical treatment; Cryo electron microscopy; Cryo electron tomography; Cryoelectron Microscopy - methods; Cytomegalovirus - chemistry; Cytomegalovirus - ultrastructure; Electron Microscope Tomography - methods; Human cytomegalovirus; Humans; Models, Molecular; pp150; Protein Conformation; Single-particle analysis; Tegument proteins; pp150; Chemical treatment; Cryo electron tomography; Cryo electron microscopy; Tegument proteins; Single-particle analysis; Human cytomegalovirus
• ISSN: 1047-8477; 1095-8657
• DOI: 10.1016/j.jsb.2011.03.006

Human cytomegalovirus (HCMV) is the most genetically and structurally complex human herpesvirus and is composed of an envelope, a tegument, and a dsDNA-containing capsid. HCMV tegument plays essential roles in HCMV infection and assembly. Using cryo electron tomography (cryoET), here we show that HCMV tegument compartment can be divided into two sub-compartments: an inner and an outer tegument. The inner tegument consists of densely-packed proteins surrounding the capsid. The outer tegument contains those components that are loosely packed in the space between the inner tegument and the pleomorphic glycoprotein-containing envelope. To systematically characterize the inner tegument proteins interacting with the capsid, we used chemical treatment to strip off the entire envelope and most tegument proteins to obtain a tegumented capsid with inner tegument proteins. SDS–polyacrylamide gel electrophoresis analyses show that only two tegument proteins, UL32-encoded pp150 and UL48-encoded high molecular weight protein (HMWP), remains unchanged in their abundance in the tegumented capsids as compared to their abundance in the intact particles. Three-dimensional reconstructions by single particle cryo electron microscopy (cryoEM) reveal that the net-like layer of icosahedrally-ordered tegument densities are also the same in the tegumented capsid and in the intact particles. CryoET reconstruction of the tegumented capsid labeled with an anti-pp150 antibody is consistent with the biochemical and cryoEM data in localizing pp150 within the ordered tegument. Taken together, these results suggest that pp150, a betaherpesvirus-specific tegument protein, is a constituent of the net-like layer of icosahedrally-ordered capsid-bound tegument densities, a structure lacking similarities in alpha and gammaherpesviruses.