Herpes Simplex Virus Type 1 (HSV-1) Uracil-DNA Glycosylase: Functional Expression in Escherichia coli, Biochemical Characterization, and Selective Inhibition by 6-( p-n-Octylanilino)Uracil

• Published in: Virology (New York, N.Y.) N.Y.), 1995-08, Vol.211 (1), p.307-311
• Main Authors: Argnani, R., Focher, F., Zucchini, S., Verri, A., Wright, G.E., Spadari, S., Manservigi, R.
• Format: Article
• Language: English
• Subjects: Base Sequence; Cloning, Molecular; Codon; DNA Glycosylases; DNA Primers; DNA Repair; Electrophoresis, Polyacrylamide Gel; Escherichia coli; Genes, Viral; herpes simplex virus 1; Herpesvirus 1, Human - enzymology; Herpesvirus 1, Human - genetics; Kinetics; Molecular Sequence Data; Molecular Weight; N-Glycosyl Hydrolases - antagonists & inhibitors; N-Glycosyl Hydrolases - isolation & purification; N-Glycosyl Hydrolases - metabolism; Polymerase Chain Reaction; Recombinant Proteins - antagonists & inhibitors; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; Restriction Mapping; Uracil - analogs & derivatives; Uracil - pharmacology; Uracil-DNA Glycosidase
• ISSN: 0042-6822; 1096-0341
• DOI: 10.1006/viro.1995.1406

The Herpes simplex virus type 1 (HSV-1) uracil-DNA glycosylase (UDG) is encoded by the UL2 gene. The translation from the first putative start codon of UL2 predicts a polypeptide of 334 residues, while the translation from the second start codon predicts a polypeptide of 244 residues. We have cloned and expressed the two forms of UDG, by means of the prokaryotic expression vector pMAL-c2, and both of them were enzymatically active. Furthermore, the enzymatic properties of the recombinant UDGs and of the enzyme purified from HSV-1-infected cells were similar. The two UDG polypeptides have molecular weights of 27 and 37 kDa, respectively. The 37-kDa form of recombinant UDG is consistent with the reported molecular mass of 37 kDa for the enzyme purified from HSV-1-infected cells. Both recombinant UDGs were as sensitive as UDG purified from HSV-1-infected cells to 6-( p-n-octylanilino)uracil, the most potent of a series of uracil analogs that inhibit the viral enzyme.