Sequence analysis of the bovine coronavirus nucleocapsid and matrix protein genes

The 3' end of the turkey coronavirus (TCV) genome and the gene encoding the nucleocapsid protein (N) were cloned and sequenced. The gene encoding the membrane protein (M) was obtained by cloning a polymerase chain reaction (PCR)-amplified fragment obtained using bovine coronavirus (BCV)-specific primers. Furthermore, five TCV DNA fragments, obtained by PCR on RNA from clinical specimens and corresponding to either the N terminus of the M protein or the complete M protein were also cloned and sequenced. The sequence revealed a 3' non-coding region of 291 bases, an open reading frame (ORF) encoding the N protein with a predicted size of 448 amino acids, or an Mr of 49K, and an ORF encoding the M protein with a predicted size of 230 amino acids and an Mr of 26K. A third ORF, encoding a hypothetical protein of 207 amino acids with an Mr of 23K was found within the N gene sequence. The amino acid sequences of both the N and M proteins were more than 99% similar to those published for BCV. Extensive similarity was also observed between the amino acid sequences of the TCV N protein and those of murine hepatitis virus (MHV) (70%) and human respiratory coronavirus strain OC43 (HCV-OC43) (98%) and between the amino acid sequences of the predicted M proteins of TCV and MHV (86 %). Such striking identity suggests that BCV, TCV and HCV-OC43 must have diverged from each other only recently. A potential N-glycosylation site was found at the N terminus of the TCV M protein and is situated at the same location in BCV, MHV and transmissible gastroenteritis virus.

Section: Sequence Analysis Of Tcv-minnesota Cdna Clonesmentioning

confidence: 58%

Sequence analysis of the turkey enteric coronavirus nucleocapsid and membrane protein genes: a close genomic relationship with bovine coronavirus

Verbeek¹,

Tijssen²

1991

“…One of the best characterized viral homodimeric glycoproteins is the H-neuraminidase protein of paramyxoviruses which happens also to exist as a homotetramer (Markwell & Fox, 1980). No data to date, however, suggest that multimeric forms other than the dimeric form exist for BCV H. One outstanding feature of BCV H is its high degree of glycosylation; 35~ of its mass is carbohydrate which compares with only 17 ~ for the peplomer protein and 13 ~ for the matrix protein (Hogue, 1986;Lapps et al, 1987). Few other viral glycoproteins are as highly glycosylated.…”

Section: Discussionmentioning

confidence: 99%

“…These data suggest that H is synthesized as separate subunits which undergo rapid disulphide linkage and glycosylation. To characterize this mode of synthesis more fully, the size of the unglycosylated protein and the mode of glycosylation were examined using an inhibitor of asparagine ( inhibitor of N-linked glycosylation (Takasuki et al, 1975), inhibited the glycosylation of the peplomeric protein and H, but not the matrix protein (Hogue, 1986;Lapps et al, 1987). Tunicamycin was therefore used in widely differing concentrations (0.5 to 10 gg/ml) and its effects on intracellular H were examined by immunoblotting.…”

Section: The 140k H Protein Is Assembled Jrom Separately Synthesized mentioning

confidence: 99%

Synthesis and Processing of the Bovine Enteric Coronavirus Haemagglutinin Protein

Hogue¹,

Kienzle²,

Brian³

1989

SUMMARYThe haemagglutinin molecule on the bovine enteric coronavirus has been identified as a glycoprotein of 140K composed of disulphide-linked subunits of 65K. In this study, we have shown the subunits to be identical by demonstrating an unambiguous aminoterminal amino acid sequence. The unglycosylated subunit was found to have an M r of 42.5K and to undergo rapid disulphide linkage and glycosylation. Glycosylation was found to be of the asparagine-linked type and some of the oligosaccharides underwent processing to complex forms. Studies with inhibitors of glycosylation suggested that a processing of the haemagglutinin oligosaccharide takes place on the virion whilst it is in the Golgi apparatus. Each haemagglutinin subunit on the mature virion was estimated to possess six or seven carbohydrate chains of either the high-mannose or hybrid type, and three or four chains of the complex type.

“…More than 10 coronavirus N genes have now been sequenced, including those of HCV 229E, TGEV, PRCV, FIPV, CCV, murine hepatitis virus (MHV), HCV OC43, bovine coronavirus (BCV) and infectious bronchitis virus (IBV) (Schreiber et al, 1989;Kapke & Brian, 1986;Britton et al, 1991 ;Vennema et al, 1991 ;Horsburgh et al, 1992;Parker & Masters, 1990;Kamahora et al, 1989;Lapps et al, 1987;Boursnell et al, 1985). Many of these viruses have been sequenced by more than one group; the references quoted here were those used in the sequence analyses shown in Fig.…”

Section: Discussionmentioning

confidence: 99%

“…Several coronaviruses possess a second ORF enclosed within the N gene, but in another reading frame. BCV has a 621 nt ORF in this position, which has recently been shown to express a protein of M r 23K in virusinfected cells and has therefore been called the internal (I) gene (Lapps et al, 1987;Senanayake et al, 1992). Amongst the coronaviruses antigenically related to BCV, four MHV strains possess a similar 621 nt ORF (Parker & Masters, 1990), whereas in MHV JHM and HCV OC43 two and three stop codons, respectively, reduce the size of the ORF (Skinner & Siddell, 1983;.…”

Section: Discussionmentioning

confidence: 99%

Sequence determination of the nucleocapsid protein gene of the porcine epidemic diarrhoea virus confirms that this virus is a coronavirus related to human coronavirus 229E and porcine transmissible gastroenteritis virus

Bridgen

Duarte

Tobler

et al. 1993

The nucleotide sequence of 1.7 kbp cDNA, comprising the region nearest the 3' end of the genome of the porcine epidemic diarrhoea virus (PEDV), has been independently determined for two European isolates of PEDV. Almost identical results were obtained for the two isolates, which were derived from cases of PEDV infection in Belgium and Britain in 1977 and 1987, respectively. The sequences contained a 1323 nucleotide (nt) open reading frame (ORF), which showed moderate identity to the nucleocapsid (N) gene of other coronaviruses. The greatest similarity at both the nucleic acid and protein levels was to the human coronavirus 229E.The PEDV N gene was, however, notably larger than that of the human 229E and porcine transmissible gastroenteritis viruses. This reflects the presence of a putative insertion of approximately 135 nt located towards the middle of the N gene. A second 336 nt ORF, which might encode a leucine-rich protein similar to, but shorter than, the bovine coronavirus internal protein was found within the PEDV N gene. Several RNA motifs typical of coronaviruses were also observed. These results confirm the earlier provisional classification of PEDV as a coronavirus.