title: Complex formation between influenza virus polymerase proteins expressed in Xenopus oocytes.
abstract: All three influenza virus polymerase (P) proteins were expressed in Xenopus oocytes from microinjected in vitro transcribed mRNA analogs, with yields of up to 100 ng per oocyte. To examine the functional state of the Xenopus-expressed P proteins, the polypeptides were tested for their ability to form stable complexes with each other. As seen in virus-infected cells, all three P proteins associated into an immunoprecipitable complex, suggesting that the system has considerable promise for the reconstruction of an active influenza RNA polymerase. Examination of the ability of paired combinations of the P proteins to associate indicated that PB1 contained independent binding sites for PB2 and PA, and so probably formed the backbone of the complex. Sedimentation analysis of free and complexed P proteins indicated that PB1 and PB2 did not exist as free monomers, and that similarly, complexes of all three P proteins did not simply consist of one copy of each protein. The heterodisperse sedimentation rate seen for complexes of all three P proteins did not appear to result from their binding to RNA, suggesting the incorporation of additional polypeptides in the polymerase complex.
authors.affiliation: Department of Pathology, University of Cambridge.
authors.forename: P
authors.initials: P
authors.lastname: Digard
authors.forename: V C
authors.initials: VC
authors.lastname: Blok
authors.forename: S C
authors.initials: SC
authors.lastname: Inglis
mesh_terms: D000818:Animals; D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000276:immunology / Q000502:physiology*; D009980:Influenza A virus / Q000201:enzymology* / Q000235:genetics; D046911:Macromolecular Substances; D008845:Microinjections; D008970:Molecular Weight; D009865:Oocytes; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D014982:Xenopus laevis
chemical_list: D046911:Macromolecular Substances; D012333:RNA, Messenger; D012321:DNA-Directed RNA Polymerases