Sequence and translation of the murine coronavirus 5'-end genomic RNA reveals the N-terminal structure of the putative RNA polymerase.
A 28-kilodalton protein has been suggested to be the amino-terminal protein cleavage product of the putative coronavirus RNA polymerase (gene A) (M.R. Denison and S. Perlman, Virology 157:565-568, 1987). To elucidate the structure and mechanism of synthesis of this protein, the nucleotide sequence of the 5' 2.0 kilobases of the coronavirus mouse hepatitis virus strain JHM genome was determined. This sequence contains a single, long open reading frame and predicts a highly basic amino-terminal region. Cell-free translation of RNAs transcribed in vitro from DNAs containing gene A sequences in pT7 vectors yielded proteins initiated from the 5'-most optimal initiation codon at position 215 from the 5' end of the genome. The sequence preceding this initiation codon predicts the presence of a stable hairpin loop structure. The presence of an RNA secondary structure at the 5' end of the RNA genome is supported by the observation that gene A sequences were more efficiently translated in vitro when upstream noncoding sequences were removed. By comparing the translation products of virion genomic RNA and in vitro transcribed RNAs, we established that our clones encompassing the 5'-end mouse hepatitis virus genomic RNA encode the 28-kilodalton N-terminal cleavage product of the gene A protein. Possible cleavage sites for this protein are proposed.
Soe
L H
LH
Department of Microbiology, University of Southern California School of Medicine, Los Angeles 90033.
Shieh
C K
CK
Baker
S C
SC
Chang
M F
MF
Lai
M M
MM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003062:Codon / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D005814:Genes, Viral; D008969:Molecular Sequence Data; D006517:Murine hepatitis virus / Q000235:genetics*; D010449:Peptide Mapping; D014176:Protein Biosynthesis; D012367:RNA, Viral / Q000235:genetics*; D014158:Transcription, Genetic
D003062:Codon; D012367:RNA, Viral; D012321:DNA-Directed RNA Polymerases