CryoEM and mutagenesis reveal that the smallest capsid protein cements and stabilizes Kaposi's sarcoma-associated herpesvirus capsid.

Abstract

With just one eighth the size of the major capsid protein (MCP), the smallest capsid protein (SCP) of human tumor herpesviruses--Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV)--is vital to capsid assembly, yet its mechanism of action is unknown. Here, by cryoEM of KSHV at 6-Å resolution, we show that SCP forms a crown on each hexon and uses a kinked helix to cross-link neighboring MCP subunits. SCP-null mutation decreased viral titer by 1,000 times and impaired but did not fully abolish capsid assembly, indicating an important but nonessential role of SCP. By truncating the C-terminal half of SCP and performing cryoEM reconstruction, we demonstrate that SCP's N-terminal half is responsible for the observed structure and function whereas the C-terminal half is flexible and dispensable. Serial truncations further highlight the critical importance of the N-terminal 10 aa, and cryoEM reconstruction of the one with six residues truncated localizes the N terminus of SCP in the cryoEM density map and enables us to construct a pseudoatomic model of SCP. Fitting of this SCP model and a homology model for the MCP upper domain into the cryoEM map reveals that SCP binds MCP largely via hydrophobic interactions and the kinked helix of SCP bridges over neighboring MCPs to form noncovalent cross-links. These data support a mechanistic model that tumor herpesvirus SCP reinforces the capsid for genome packaging, thus acting as a cementing protein similar to those found in many bacteriophages.

• Authors: Xinghong Dai, Danyang Gong, Yuchen Xiao, Ting-Ting Wu, Ren Sun, Z Hong Zhou
• Journal: Proceedings of the National Academy of Sciences of the United States of America (Proc Natl Acad Sci U S A) Vol. 112 Issue 7 Pg. E649-56 (Feb 17 2015) ISSN: 1091-6490 [Electronic] United States
• PMID: 25646489 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, Non-P.H.S.)

Chemical References

• DNA Primers

Topics

• Amino Acid Sequence
• Base Sequence
• Capsid (metabolism, ultrastructure)
• Cell Line
• Cryoelectron Microscopy (methods)
• DNA Primers
• Herpesvirus 8, Human (genetics, metabolism, ultrastructure)
• Humans
• Molecular Sequence Data
• Mutagenesis
• Protein Conformation
• Sequence Homology, Amino Acid