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...dicated that the 290 kDa protein is the precursor of the 50 kDa and 240 kDa proteins (Denison et al., 1992). The experimental data also suggested that p65 is adjacent to the carboxyl terminus of p28 (=-=Denison et al., 1995-=-). Subsequently, a second PL1pro cleavage site was identified (Bonilla et al., 1995) and characterized by amino acid sequence analysis (Bonilla et al., 1997). This cleavage occurs at the Ala-832 � Gly...

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...is (75). Reactions of specific antisera raised against different regions of MHV POL1awith potential cleavage products with apparent molecular weights of 65, 50, 240, and 290 kDa in MHV-infected cells =-=(80,81)-=- showed that processing of the N-terminus of POL1a involves multiple cleavage events. p65 is thought to be immediately adjacent to p28 (81,82). Gao et al. (82) reported that p65 of MHV strain JHM is g...

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...r the synthesis of the viral RNA-dependent RNA polymerase (RdRp) polyprotein via the cap-dependent translation mechanism, which is subsequently processed into 16 nonstructural peptides (nsps 1 to 16) =-=(2, 3, 5, 12, 13, 21, 30, 31, 37, 42, 44, 47)-=-. The polymerase complex in turn initiates viral RNA synthesis, including genomic RNA replication and subgenomic mRNA synthesis. In addition to the RdRp, a number of host cell proteins such as heterog...

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... proteinases, papain-like proteinase (PLP) 1, PLP2, and 3C-like proteinase (3CLpro) to produce intermediate and mature replicase proteins (Baker et al., 1989; Bonilla et al., 1997; Bost et al., 2000; =-=Denison et al., 1995-=-; Graham and Denison, 2006; Kanjanahaluethai and Available online at www.sciencedirect.com Virology 375 (2008) 118–129 www.elsevier.com/locate/yviro ⁎ Corresponding author. Department of Microbiology ...

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...d from pp1a/1ab weresdetected in vivo. Two large processed products (approximately 240 and 290 kDa in size) of thesORF1a-derived polyprotein of murine hepatitis virus (MHV) were successfully detected =-=[41; 42]-=-. Ins1998, several cleavage sites of MHV were predicted and verified as well and a 22 kDa proteinsfragment of polyprotein 1a (ORF1a) was observed in MHV infected cells using a specific antibodys[43]. ...

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...ollowing high-salt translational synchronization to ensure that detected proteins were de novo translation products rather than proteins that were partially translated prior to addition of radiolabel =-=(6, 23)-=-. The proteins were immunoprecipitated with B4 antiserum. In infected cells, p1a22 and the 200-kDa protein were first observed at 60 min of labeling and accumulated without detectable intermediates (F...

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...critical for embedding the replicase complex into cellular membranes. p150 itself is extensively processed by MHV 3CLpro (37). In addition, the region upstream of p150 is processed by the PLP1 domain =-=(2, 3, 5, 6, 8, 16)-=-. Therefore, at least three distinct proteinase activities, PLP1, PLP2, and 3CLpro, are required for the maturation of the MHV replicase complex. Previously, we identified the p150 intermediate as a p...

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...oronaviruses, with the exception of proteins expressed from the amino terminus. Specifically, the group 2 coronaviruses cleave unique 28- and 65-kDa aminoterminal proteins (p28 and p65, respectively) =-=(10, 12)-=-. The functions of p28 and p65 are not known, and neither protein is expressed during the infectious cycle of the group 1 and 3 * Corresponding author. Mailing address: Lamb Center for Pediatric Resea...

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...ing bovine coronavirus and mouse hepatitis virus (MHV), PLP1 has been shown in vitro to cleave an amino-terminal 28-kDa protein and a 65-kDa protein, referred to as p28 and p65, respectively (Fig. 1) =-=(1, 2, 7, 8, 10, 14)-=-. In contrast, SARS-CoV, as well as the avian coronaviruses (group 3), such as infectious bronchitis virus (IBV), only express active PLP2, which is responsible for cleavage of amino-terminal proteins...

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...ll of the three ORF1a-encoded proteinases are believed to process the replicase polyprotein into mature polypeptides. Some of the ORF1a-encoded proteins expressed intracellularly have been identified =-=(12, 13, 23, 34)-=-. The first virus-encoded nonstructural polypeptide identified was the Nterminal p28, which is rapidly processed from the ORF1a polyprotein. Because p28 is processed in vitro by the PLP-1, it is belie...