Characterization and nuclear localization of the fiber protein encoded by the late region 7 of bovine adenovirus type 3

Characterization and nuclear localization of the fiber protein encoded by the late region 7 of... To identify the protein encoded by the L7 region of bovine adenovirus-3 (BAdV-3), specific antisera were raised by immunizing rabbits with bacterial fusion proteins encoding the N-terminus or C-terminus of the BAdV-3 fiber protein. Immunoprecipitation and Western blot analysis confirmed that the fiber is expressed as a 102 kDa glycoprotein, which is localized to the nucleus of infected cells. To identify the nuclear localization signals (NLS), BAdV-3 fiber deletion mutants and GFP/β-galactosidase fusion proteins were expressed in transfected cells, and subcellular localization was visualized by immunofluorescence microscopy. Analysis of deletion mutants localized the NLS to the N-terminal 41 amino acids. Analysis of the N-terminal 41 amino acids identified a cluster of basic residues between amino acid 14 and 20. Substitution of the basic residues ( 16 KAKR 19 ) with acidic residues ( 16 EAEE 19 ) resulted in the accumulation of fiber in the cytoplasm. However, 16 KAKR 19 or 12 VYPYKAKRPNI 22 were not sufficient for efficient transport of a cytoplasmic protein GFP/β-galactosidase to the nucleus. The recombinant BAdV-3 expressing mutant fiber containing 16 EAEE 19 instead of 16 KAKR 19 was unable to replicate efficiently in Madin-Darby bovine kidney cells, suggesting that the NLS of fiber carries out important in vivo functions. Archives of Virology Springer Journals

Characterization and nuclear localization of the fiber protein encoded by the late region 7 of bovine adenovirus type 3

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