Sequence analysis of divergent canine coronavirus strains present in a UK dog population.

Forty faecal samples were tested by RT-PCR using coronavirus consensus primers to determine faecal shedding of canine coronavirus (CCoV) and canine respiratory coronavirus (CRCoV) in a dog population housed at a rescue centre. Seven samples were positive for CCoV while all samples were negative for CRCoV. Sequence analysis of five CCoV strains showed a high similarity with transmissible gastroenteritis virus (TGEV) at the N-terminus of the spike protein. All strains contained an open reading frame for the nonstructural protein 7b, which is not present in TGEV, indicating that the strains were related to the previously described CCoV strain UCD-1. Two samples contained CCoV strains with 5' spike sequences most similar to type II CCoV while one sample was found to contain type I CCoV. Primers directed to the N gene allowed specific detection of all CCoV strains analysed in this study. This investigation shows that CCoV strains containing spike proteins similar to TGEV are present in the UK dog population. PCR primers directed to conserved regions of the CCoV genome are recommended for detection of CCoV in clinical samples due to high genetic variability.

Small envelope protein E of SARS: cloning, expression, purification, CD determination, and bioinformatics analysis.

AIM: To obtain the pure sample of SARS small envelope E protein (SARS E protein), study its properties and analyze its possible functions. METHODS: The plasmid of SARS E protein was constructed by the polymerase chain reaction (PCR), and the protein was expressed in the E coli strain. The secondary structure feature of the protein was determined by circular dichroism (CD) technique. The possible functions of this protein were annotated by bioinformatics methods, and its possible three-dimensional model was constructed by molecular modeling. RESULTS: The pure sample of SARS E protein was obtained. The secondary structure feature derived from CD determination is similar to that from the secondary structure prediction. Bioinformatics analysis indicated that the key residues of SARS E protein were much conserved compared to the E proteins of other coronaviruses. In particular, the primary amino acid sequence of SARS E protein is much more similar to that of murine hepatitis virus (MHV) and other mammal coronaviruses. The transmembrane (TM) segment of the SARS E protein is relatively more conserved in the whole protein than other regions. CONCLUSION: The success of expressing the SARS E protein is a good starting point for investigating the structure and functions of this protein and SARS coronavirus itself as well. The SARS E protein may fold in water solution in a similar way as it in membrane-water mixed environment. It is possible that beta-sheet I of the SARS E protein interacts with the membrane surface via hydrogen bonding, this beta-sheet may uncoil to a random structure in water solution.