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Journal of Virology, December 2001, p. 12228-12240, Vol. 75, No. 24
Department of Molecular and Cell
Biology1 and Proteomics
Laboratory,2 Centro Nacional de
Biotecnología, CSIC, Campus Universidad Autónoma,
Cantoblanco, 28049 Madrid, Spain, and Unité de
Virologie Immunologie Moléculaires, INRA, 78350 Jouy-en-Josas,
France3
Received 6 July 2001/Accepted 6 September 2001
The difference in membrane (M) protein compositions
between the transmissible gastroenteritis coronavirus (TGEV) virion and the core has been studied. The TGEV M protein adopts two topologies in
the virus envelope, a Nexo-Cendo topology (with the amino terminus exposed to the virus surface and the carboxy terminus inside the virus
particle) and a Nexo-Cexo topology (with both the amino and carboxy
termini exposed to the virion surface). The existence of a population
of M molecules adopting a Nexo-Cexo topology in the virion envelope was
demonstrated by (i) immunopurification of 35S-labeled TGEV
virions using monoclonal antibodies (MAbs) specific for the M protein
carboxy terminus (this immunopurification was inhibited only by
deletion mutant M proteins that maintained an intact carboxy terminus),
(ii) direct binding of M-specific MAbs to the virus surface, and (iii)
mass spectrometry analysis of peptides released from trypsin-treated
virions. Two-thirds of the total number of M protein molecules found in
the virion were associated with the cores, and one-third was lost
during core purification. MAbs specific for the M protein carboxy
terminus were bound to native virions through the M protein in a
Nexo-Cexo conformation, and these molecules were removed when the virus envelope was disrupted with NP-40 during virus core purification. All
of the M protein was susceptible to N-glycosidase F treatment of the
native virions, which indicates that all the M protein molecules are
exposed to the virus surface. Cores purified from glycosidase-treated
virions included M protein molecules that completely or partially lost
the carbohydrate moiety, which strongly suggests that the M
protein found in the cores was also exposed in the virus envelope and
was not present exclusively in the virus interior. A TGEV
virion structure integrating all the data is proposed. According to
this working model, the TGEV virion consists of an internal core, made
of the nucleocapsid and the carboxy terminus of the M
protein, and the envelope, containing the spike (S) protein, the
envelope (E) protein, and the M protein in two conformations. The two-thirds of the molecules that are in a Nexo-Cendo conformation (with their carboxy termini embedded within the virus core) interact with the internal core, and the remaining third of
the molecules, whose carboxy termini are in a Nexo-Cexo conformation, are lost during virus core purification.
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.24.12228-12240.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Organization of Two Transmissible Gastroenteritis
Coronavirus Membrane Protein Topologies within the Virion and
Core
*
Corresponding author. Mailing address: Department of
Molecular and Cell Biology, Centro Nacional de Biotechnología,
CSIC, Campus Universidad Autónoma, Cantoblanco, 28049 Madrid,
Spain. Phone: 34-91-585-4555. Fax: 34-91-585-4915. E-mail:
L.Enjuanes{at}cnb.uam.es.
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