RNA-stimulated NTPase activity associated with the p80 protein of the pestivirus bovine viral diarrhea virus.
The genomic RNA of pestiviruses contains a single large open frame coding for virion structural proteins and viral nonstructural polypeptides. Based on the presence of specific amino acid sequence motifs, pestivirus nonstructural protein p80 was predicted to be both a serine-type proteinase and a nucleoside triphosphatase (NTPase)/RNA helicase. We previously demonstrated p80 possesses the former activity (Wisherchen and Collett, Virology 184, 341-350, 1991). Here, we provide experimental evidence that this protein is also an RNA-stimulated NTPase. Employing immunoaffinity chromatography, we partially purified a p80 protein analog (p87) from recombinant baculovirus-infected insect cells. We show this preparation contained a specific NTPase activity. This activity was not found in material similarly purified from lysates of baculovirus-infected insect cells not expressing the p87 protein. That the NTPase activity was associated with the p87 polypeptide was demonstrated in two ways. First, the NTPase activity was shown to be completely inhibited by monoclonal antibodies specific to the p80 polypeptide, but was unaffected by monoclonal antibodies to unrelated antigens. Second, radiolabeled ATP could be specially cross-linked to the p87 polypeptide. NTP hydrolysis by the p87 protein was stimulated by the presence of particular single-strand RNA molecules. Initial enzymologic characterization of the pestivirus p80 NTPase is presented, and the presumptive role of this activity in pestivirus replication is discussed.
Medimmune, Inc., Gaithersburg, Maryland 20878.
J K
JK
Tamura
P
P
Warrener
M S
MS
Collett
D000818:Animals; D016367:Baculoviridae; D001908:Diarrhea Viruses, Bovine Viral / Q000737:chemistry / Q000201:enzymology*; D007915:Lepidoptera; D043583:Nucleoside-Triphosphatase; D010744:Phosphoric Monoester Hydrolases / Q000302:isolation & purification / Q000378:metabolism*; D012367:RNA, Viral / Q000502:physiology; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000302:isolation & purification / Q000378:metabolism*
D012367:RNA, Viral; D011994:Recombinant Proteins; D017361:Viral Nonstructural Proteins; D010744:Phosphoric Monoester Hydrolases; D043583:Nucleoside-Triphosphatase