Proteolytic mapping of the coronavirus infectious bronchitis virus 1b polyprotein: evidence for the presence of four cleavage sites of the 3C-like proteinase and identification of two novel cleavage products.
We have previously reported that the 3C-like proteinase of the coronavirus infectious bronchitis virus (IBV) is responsible for processing of the 1a and 1a/1b polyproteins to three mature products of 24, 10, and 100 kDa (Liu et al., 1994, 1997; Ng and Liu, 1998). The C-terminal cleavage site of the 100-kDa protein was defined to be the Q891(1b)-S892(1b) dipeptide bond encoded by nucleotides 15,129 to 15,134 (Liu and Brown, 1995). In this report, other cleavage sites of the 3C-like proteinase in the polyprotein encoded by the ORF 1b region were mapped by coexpression, deletion, and site-directed mutagenesis studies. Using two ORF 1b-specific antisera, V58 and V17, three more Q-S(G) dipeptide bonds, encoded by nucleotides 16,929 to 16,934, 18,492 to 18,497, and 19,506 to 19,511, respectively, were demonstrated to be the cleavage sites of the 3C-like proteinase. Cleavage at these four positions would result in the release of four mature products with molecular masses of approximately 68, 58, 39, and 35 kDa. Among them, the 39- and 35-kDa proteins were specifically identified in IBV-infected cells. Taken together with the 100-kDa protein previously identified, these results suggest that the ORF 1b region of IBV mRNA1 may be able to encode five mature products.
Liu
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Institute of Molecular Agrobiology, National University of Singapore, Singapore. liudx@ima.org.sg
Shen
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S
Xu
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Wang
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D000818:Animals; D001665:Binding Sites; D002522:Chlorocebus aethiops; D000086782:Coronavirus 3C Proteases; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D001351:Infectious bronchitis virus / Q000201:enzymology*; D016297:Mutagenesis, Site-Directed; D016366:Open Reading Frames; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D014709:Vero Cells; D014764:Viral Proteins / Q000378:metabolism*
D011498:Protein Precursors; D014764:Viral Proteins; C087637:gene 1 protein, Coronavirus; D003546:Cysteine Endopeptidases; C000706208:3C-like proteinase, Avian infectious bronchitis virus; D000086782:Coronavirus 3C Proteases