We have reported previously that the herpes simplex virus type 1 (HSV-1) thymidine kinase (TK) fused with green fluorescent protein (GFP) is localized in the nucleus of HSV-1 TK-GFP gene-transfected cells (Degrève et al. (1998) J. Virol. 72, 9535-9543). Deletion of the N-terminal 34 amino acids or selective mutation of the nonapeptide (25)RRTALRPRR(33), located in the N-terminal region of HSV-1 TK, resulted in the loss of the specific nuclear localization of HSV-1 TK. Utilizing information on the crystallographic structure of HSV-1 TK, we have now identified three additional putative nuclear localization signals and evaluated their potential role in the nuclear trafficking of HSV-1 TK by site-directed mutagenesis. We found that the sites containing the amino acids R236-R237 and K317-R318 are absolutely required for specific nuclear targeting of HSV-1 TK. The K317-R318 region, located at the interface between the two monomers in the dimeric HSV-1 TK structure, could act as a nuclear localization signal for monomeric HSV-1 TK. Alternatively, crystallographic data indicate that R318 might be essential for the formation of the TK dimer, and therefore it is required if HSV-1 TK is transported as a dimer.

Original publication

DOI

10.1006/bbrc.1999.1485

Type

Journal article

Journal

Biochem Biophys Res Commun

Publication Date

22/10/1999

Volume

264

Pages

338 - 342

Keywords

Amino Acid Sequence, Binding Sites, Cell Nucleus, Cytosol, Green Fluorescent Proteins, Herpesvirus 1, Human, Humans, Luminescent Proteins, Molecular Sequence Data, Mutagenesis, Site-Directed, Mutation, Plasmids, Recombinant Fusion Proteins, Thymidine Kinase, Transfection, Tumor Cells, Cultured