Very little is known about the astrovirus coat protein precursor and how it assembles into particles. In an attempt to understand what regions of this protein might contribute to particle assembly and receptor binding, we have utilized a structural prediction program to analyze the coat protein amino acid sequence. This analysis has led to predictions about what domains of the coat protein are necessary for protein assembly, as well as for host cell tropism. Except for within the Astroviridae
, the astrovirus coat protein exhibits no significant sequence identity with any known viral capsid protein. As illustrated in above, the coat protein of both the Mamastrovirus
has been divided into conserved (residues 1-415) and variable (residues 416-end) regions. Because of the lack of sequence identity seen in the variable region of the astrovirus coat protein (65
) and based on the fact that neutralizing antibodies map to this area, (3
) it has been proposed that the variable region is exposed on the surface of the particle and is a determinant for cellular tropism. In light of the conserved nature of residues 1-415, we reasoned that this region of the coat protein may represent a core “assembly domain”, a building block for capsid assembly. In support of this, a threonine residue at amino acid 227 of the HAstV-1 coat protein was demonstrated to be required for proper particle assembly (38
). Mutation of this residue resulted in the absence of visible particles within the cytoplasm of CaCo-2 cells.
We used three-dimensional position-specific scoring matrix (3D-PSSM, version 2.6.0) to investigate whether conserved and variable regions of the coat protein assume a recognizable protein-folding motif (26
). 3D-PSSM combines knowledge of three-dimensional structures with secondary-structure matching and solvation potentials to recognize protein folds when sequence homologies are remote. A number of parameters go into assessing confidence of the predictions of this program: Expectation Value (or Confidence Value), alignment quality and 3D molecule quality. Confidence Value scores are confirmed by checking the sequence alignment. A lack of significant gaps in the alignment and a match of the secondary structural elements are signs of good alignment. For model quality, a lack of significant deletions in the template (model) structure is important. We considered these factors to make predictions about structural information contained in the astrovirus capsid gene sequence. Previously, we applied this program to determine the domain organization of another icosahedral RNA virus that was verified by cryo-electron microscopy (58