Site-directed mutagenesis in the active site of the herpes simplex virus type 1 thymidine kinase gene.
The thymidine kinase (TK) of herpes simplex virus type 1 (HSV-1) contains three regions of homology to other ATP utilizing enzymes. We have altered one region of the protein, which seems to play an important role in phosphorylation substrates by site-directed mutagenesis. When the aspartate 162 was changed to asparagine, the enzyme lost its activity. To identify the inactive protein, expressed by a vaccinia vector in eukaryotic cells, a monospecific antiserum against a bacterial tryptophan E-HSV-1 TK fusion protein was made. These results support the suggestion that aspartate 162 is essential for the enzymatic activity.
Department of Pharmacy, Swiss Federal Institute of Technology, Zürich.
J
J
Fetzer
G
G
Folkers
I
I
Müller
G M
GM
Keil
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D005814:Genes, Viral; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011993:Recombinant Fusion Proteins / Q000235:genetics; D018139:Simplexvirus / Q000201:enzymology / Q000235:genetics*; D013937:Thymidine Kinase / Q000235:genetics*; D014616:Vaccinia virus / Q000235:genetics
D004279:DNA, Viral; D011993:Recombinant Fusion Proteins; D013937:Thymidine Kinase