Localization of the N-terminus of Minor Coat Protein IIIa in the Adenovirus Capsid

Overview

Journal J Mol Biol

Publisher Elsevier

Specialties Microbiology
Molecular Biology

Date 2008 Sep 13

PMID 18786542

Citations 25

Authors

Carmen San Martin

Joel N Glasgow

Anton Borovjagin

Matthew S Beatty

Elena A Kashentseva

David T Curiel

Roberto Marabini

Igor P Dmitriev

Affiliations

Soon will be listed here.

Abstract

Minor coat protein IIIa is conserved in all adenoviruses (Ads) and is required for correct viral assembly, but its precise function in capsid organization is unknown. The latest Ad capsid model proposes that IIIa is located underneath the vertex region. To obtain experimental evidence on the location of IIIa and to further define its role, we engineered the IIIa gene to encode heterologous N-terminal peptide extensions. Recombinant Ad variants with IIIa encoding six-histidine (6His) tag, 6His, and FLAG peptides, or with 6His linked to FLAG with a (Gly(4)Ser)(3) linker were rescued and analyzed for virus yield, capsid incorporation of heterologous peptides, and capsid stability. Longer extensions could not be rescued. Western blot analysis confirmed that the modified IIIa proteins were expressed in infected cells and incorporated into virions. In the Ad encoding the 6His-linker-FLAG-IIIa gene, the 6His tag was present in light particles, but not in mature virions. Immunoelectron microscopy of this virus showed that the FLAG epitope is not accessible to antibodies on the viral particles. Three-dimensional electron microscopy and difference mapping located the IIIa N-terminal extension beneath the vertex complex, wedged at the interface between the penton base and peripentonal hexons, therefore supporting the latest proposed model. The position of the IIIa N-terminus and its low tolerance for modification provide new clues for understanding the role of this minor coat protein in Ad capsid assembly and disassembly.

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