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J. Virol. doi:10.1128/JVI.00986-07
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Structural features of the scaffold interaction domain (SID) at the N-terminus of the major capsid protein (VP5) of HSV-1

Viral Oncology Program, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Department of Biology and Integrated Imaging Center, Johns Hopkins University, Baltimore, MD

^* To whom correspondence should be addressed. Email: pdesai{at}jhmi.edu.

	Abstract

Protein-protein interactions drive the assembly of the herpes simplex virus type 1 capsid. A key interaction occurs between the C-terminus of the scaffold protein and the N-terminus of the major capsid protein (VP5). Results from alanine-scanning mutagenesis of hydrophobic residues in the N-terminus of VP5 revealed seven residues (I27, L35, F39, L58, L65, L67, and L71) that reside in two predicted alpha helices (helix 1 ^22-42 and helix 2 ^58-72) that are important for this bi-molecular interaction. The goal of the present study was to further characterize the VP5 scaffold interaction domain (SID). The seven positions were substituted for L, M, V or P (I27), I, M, V or P (L35, L58, L65, L67 and L71) and H, W, Y or L (F39). Substitution to a hydrophobic side-chain did not affect the interaction with scaffold protein in yeast cells or the ability of a virus specifying the mutation from replicating in cells. The mutation to the proline side-chain abolished the interaction in all cases and was lethal for virus replication. Mutant viruses with proline substitutions in helix 1 at position 27 and 35 assembled large open capsid shells that did not attain closure. Proline substitutions in helix 2 ^58-72 at either position 59, 65 and 67 abolished the accumulation of VP5 protein and at 58 and 71 although VP5 did accumulate, capsid shells were not assembled. Thus the second scaffold interaction domain (SID2) is highly structured and this alpha helix (helix 2 ^58-72) is likely involved in capsomere-capsomere interactions during shell accretion. A conserved glycine, G59 in helix 2 ^58-72 was also mutated. G59 may act as a flexible "hinge" in helix 2 because decreasing the movement of this side chain by substitution to valine impaired capsid assembly. Thus the N-terminus of VP5 and the alpha helices embedded in this domain like the capsid shell proteins of some dsDNA phages is a key regulator of shell accretion and stabilization.

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