Identification of protease and ADP-ribose 1''-monophosphatase activities associated with transmissible gastroenteritis virus non-structural protein 3.
The replicase polyproteins, pp1a and pp1ab, of porcine Transmissible gastroenteritis virus (TGEV) have been predicted to be cleaved by viral proteases into 16 non-structural proteins (nsp). Here, enzymic activities residing in the amino-proximal region of nsp3, the largest TGEV replicase processing product, were characterized. It was shown, by in vitro translation experiments and protein sequencing, that the papain-like protease 1, PL1(pro), but not a mutant derivative containing a substitution of the presumed active-site nucleophile, Cys(1093), cleaves the nsp2|nsp3 site at (879)Gly|Gly(880). By using an antiserum raised against the pp1a/pp1ab residues 526-713, the upstream processing product, nsp2, was identified as an 85 kDa protein in TGEV-infected cells. Furthermore, PL1(pro) was confirmed to be flanked at its C terminus by a domain (called X) that mediates ADP-ribose 1''-phosphatase activity. Expression and characterization of a range of bacterially expressed forms of this enzyme suggest that the active X domain comprises pp1a/pp1ab residues Asp(1320)-Ser(1486).
Institute of Virology and Immunology, University of Würzburg, Versbacher Straße 7, 97078 Würzburg, Germany.
Ákos
Á
Putics
Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, Leiden, The Netherlands.
Alexander E
AE
Gorbalenya
Institute of Virology and Immunology, University of Würzburg, Versbacher Straße 7, 97078 Würzburg, Germany.
John
J
Ziebuhr
D000595:Amino Acid Sequence; D000818:Animals; D020134:Catalytic Domain / Q000502:physiology; D002460:Cell Line; D000086882:Coronavirus Papain-Like Proteases; D005761:Gastroenteritis, Transmissible, of Swine / Q000821:virology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010206:Papain / Q000378:metabolism; D010447:Peptide Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D005760:Transmissible gastroenteritis virus / Q000378:metabolism*; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D011994:Recombinant Proteins; D017361:Viral Nonstructural Proteins; D010744:Phosphoric Monoester Hydrolases; D010447:Peptide Hydrolases; D000086882:Coronavirus Papain-Like Proteases; D010206:Papain