IDENTIFICATION, EXPRESSION, AND PROCESSING OF AN 87-KDA POLYPEPTIDE ENCODED BY ORF 1A OF THE CORONAVIRUS INFECTIOUS-BRONCHITIS VIRUS

Nucleotide sequence analysis has shown previously that the genomic-len gth mRNA (mRNA 1) of the coronavirus infectious bronchitis virus (IBV) contains two large open reading frames (ORFs), 1a and 1b, with the po tential to encode polyproteins of approximately 441 and 300 kDa, respe ctively. We have characterized the specificity of a set of region-spec ific antisera raised against the 5'-portion of ORF la by immunoprecipi tation of in vitro-synthesized, C-terminally truncated la polypeptides and used these antisera to detect virus-specific proteins in IBV-infe cted vero cells. Two antisera, which had specificity for IBV sequences from nucleotides 710 to 2079 and 1355 to 2433, respectively, immunopr ecipitated a polypeptide of approximately 87 kDa from IBV-infected Ver o cells. In vitro translation of ORF la sequence terminating at nucleo tide 5763 did not produce this protein unless the in vitro translation products were incubated with Vero cell sin extracts prepared from eit her IBV-infected or mock-infected Vero cells, However, processing of t he 87-kDa protein was also observed when the same region was expressed in Vero cells using the vaccinia virus/T7 expression system. This obs ervation indicates that the 87-kDa polypeptide is encoded within the 5 '-most 3000 nucleotides of mRNA I and that it might be cleaved from th e la polyprotein by viral and cellular proteinases. (C) 1995 Academic Press, Inc.