Proposed three-dimensional model for the attachment protein G of respiratory syncytial virus

Abstract

Protein G of respiratory syncytial virus (RSV) is an envelope glycoprotein that is structurally very different from its counterparts (haemagglutinin-neuraminidase and haemagglutinin) in other paramyxoviruses. In this study, we put forward a model for this unique viral envelope protein. We propose that protein G of RSV contains several independently folding regions, with the ectodomain consisting of a conserved central hydrophobic region located between two polymeric mucinlike regions. The central conserved region is probably the only relatively fixed and folded part of the ectodomain of RSV-G. This central conserved region contains four conserved cysteine residues which can form two disulphide bridges. Analysis of the proteolytic digestion products of a peptide corresponding to the central conserved region showed that one of the three theoretically possible combinations of disulphide connections could be eliminated. The final disulphide bridge assignment was established by affinity measurements with peptide variants in which different disulphide connections were formed. Additionally, peptide binding studies were used to map the binding site, at the amino acid level, of a monoclonal antibody directed against the central conserved region. These studies indicated the level of surface exposure of the amino acid side-chains. The surface exposure agreed with the structural model. The proteolytic digestion, the peptide binding studies and the affinity measurements with structural peptide variants support a structural model with disulphide connections that correspond to a structural motif called a cystine noose. This model provides a structural explanation for the location and molecular details of important antigenic sites.

Validation of Synthetic Peptide Enzyme Immunoassays in Differentiating Two Subgroups of Ruminant Respiratory Syncytial Virus

“…15 The carbohydrates in the mucin-like regions are extensively O-linked, forming rigid stalk-like structures that act in extending the functional domain to a particular height above the level of the cell surface. 12,15 The location of the peptides (used in this study) in the 3-dimensional model makes them antigenically attractive to the immune system and functionally significant in virus-cell attachment. The selection of negative threshold level (negative mean plus 3 SDs) was based on the distribution (data not shown) of ELISA results in 2 different populations (cattle without specific antibody and cattle with specific antibody).…”

“…Both viruses share a common peptide motif, located on a similar region of each respective G-glycoprotein, which contains 4 conserved cysteine residues comprising an antigenic cysteine loop. 15 The proposed 3-dimensional model for the RSV G protein places the 4 conserved cysteine residues into a hydrophobic globular region bordered by 2 hydrophilic mucin-like regions. 15 The carbohydrates in the mucin-like regions are extensively O-linked, forming rigid stalk-like structures that act in extending the functional domain to a particular height above the level of the cell surface.…”

“…15 The proposed 3-dimensional model for the RSV G protein places the 4 conserved cysteine residues into a hydrophobic globular region bordered by 2 hydrophilic mucin-like regions. 15 The carbohydrates in the mucin-like regions are extensively O-linked, forming rigid stalk-like structures that act in extending the functional domain to a particular height above the level of the cell surface. 12,15 The location of the peptides (used in this study) in the 3-dimensional model makes them antigenically attractive to the immune system and functionally significant in virus-cell attachment.…”

“…2 The proposed 3-dimensional model for the Gglycoprotein of BRSV contains a hydrophobic globular region (residues 158-189) bordered by 2 hydrophilic densely carbohydrate-coated mucin-like regions. 15 The G-glycoprotein gene from an ORSV strain has been cloned and sequenced and had greater homology to the BRSV gene (and its encoded protein) than to the HRSV gene (and its encoded protein). 2,4 However, the degree of base sequence differences between BRSV and ORSV suggests that 2 subgroups of an ungulate RSV exist.…”

“…2,4 However, the degree of base sequence differences between BRSV and ORSV suggests that 2 subgroups of an ungulate RSV exist. 2,15 Recent RNase protection assays, Received for publication October 26, 1999. reverse transcription polymerase chain reaction amplification studies, and a phosphoprotein profile analysis support the existence of 2 ruminant RSV subgroups. 1,9,17,18 Current knowledge concerning interspecies transmission of RSV strains is limited.…”