Abstract
The N-terminal S1 region of the transmissible gastroenteritis virus (TGEV) spike (S) glycoprotein contains four antigenic sites (C, B, D and A, from the N- to the C-terminal end) and is engaged in host-cell receptor recognition. The most N-terminal portion of the S1 region, which comprises antigenic sites C and B, is needed for the enteric tropism of TGEV, whereas the major antigenic site A at the C-terminal moiety is required for both respiratory and enteric cell tropism, and is engaged in recognition of the aminopeptidase N (APN) receptor. This study determined the kinetics for binding of a soluble S1 protein to the APN protein. Moreover, the S1 region of the TGEV S protein was dissected, with the aim of identifying discrete modules displaying unique antigenic sites and receptor-binding functions. Following protease treatments and mammalian cell expression methods, four modules or domains (D1-D4) were defined at the S1 region. Papain treatment identified an N-terminal domain (D1) resistant to proteolysis, whereas receptor binding defined a soluble and functional APN receptor-binding domain (D3). This domain was recognized by neutralizing antibodies belonging to the antigenic site A and therefore could be used as an immunogen for the prevention of viral infection. The organization of the four modules in the S1 region of the TGEV S glycoprotein is discussed.
Highlights
Coronaviruses (CoVs) are enveloped, positive-sense, ssRNA viruses (de Groot et al, 2008; Enjuanes et al, 2008; Masters, 2006) involved in respiratory, enteric, hepatic and neuronal infectious diseases in animals and humans that often lead to important economic losses (Perlman, 1998; Weiss & Navas-Martin, 2005)
Aiming to isolate folding modules displaying unique antigenic and functional properties linked to the S1 region of the transmissible gastroenteritis virus (TGEV) S protein, length variants were designed following the antigenic map described with TGEV escape mutants (Correa et al, 1990; Gebauer et al, 1991; Sanchez et al, 1992) (Fig. 1a)
Similar variants were engineered for the same region of the porcine respiratory CoV (PRCV) HOL87 strain (S1H and S3H) (Fig. 1a), which has 96 % sequence identity with the TGEV protein
Summary
Coronaviruses (CoVs) are enveloped, positive-sense, ssRNA viruses (de Groot et al, 2008; Enjuanes et al, 2008; Masters, 2006) involved in respiratory, enteric, hepatic and neuronal infectious diseases in animals and humans that often lead to important economic losses (Perlman, 1998; Weiss & Navas-Martin, 2005). Members of the genus Betacoronavirus such as human SARS-CoV use the human angiotensin-converting enzyme 2 (ACE2) (Li et al, 2003), whereas murine hepatitis virus (MHV) uses the cell adhesion molecule CEACAM1a (Yokomori & Lai, 1992). The existence of alternative receptors that can confer an extended tropism has been revealed for several CoVs. SARS-CoV can use liver/lymph node-specific intercellular adhesion molecule-3-grabbing non-integrin (L-SIGN) for cell attachment and entry (Jeffers et al, 2004), whereas an MHV virus strain isolated from persistently infected cells (MHV/BHK) enters the cells in a heparan sulfatedependent manner (de Haan et al, 2005; Miura et al, 2008). It has been suggested that binding of the TGEV S protein to sialic acids is responsible for its enteric tropism (Krempl et al, 1997)
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