Complex of the herpes simplex virus initiator protein UL9 with DNA as a platform for the design of a new type of antiviral drugs

• Published in: Biophysics (Oxford) Vol. 55; no. 2; pp. 204 - 214
• Main Authors: Surovaya, A. N., Grokhovsky, S. L., Gursky, Ya. G., Andronova, V. L., Arkhipova, V. S., Bazhulina, N. P., Galegov, G. A., Gursky, G. V.
• Format: Journal Article
• Language: English
• Published: Dordrecht SP MAIK Nauka/Interperiodica 01.04.2010; Springer Nature B.V
• Subjects: Binding sites; Biological and Medical Physics; Biophysics; Deoxyribonucleic acid; DNA; Escherichia coli; Herpes simplex virus 1; Herpes viruses; Herpesvirus; Molecular biology; Molecular Biophysics; Physics; Physics and Astronomy; Proteins; antiviral drugs; binding constant; replication origin; helicase of herpes simplex virus; bis-netropsins; ATPase and helicase activities
• ISSN: 0006-3509; 1555-6654
• DOI: 10.1134/S0006350910020077

The protein binding to the origin of replication of the herpes simplex virus type 1 is DNA helicase encoded by the UL9 gene of the herpes virus. The protein specifically binds to two binding sites in the viral DNA replication origins OriS or OriL. In order to determine the role of the UL9 protein in the initiation of replication and find efficient inhibitors of the UL9 activity, we have synthesized a recombinant UL9 protein expressed in E. coli cells. It was found that the recombinant UL9 protein binds to Boxes I and II in OriS and possesses DNA helicase and ATPase activities. In the complex with a fluorescent analog of ATP, two molecules of the ATP analog bind to one protein dimer molecule. It was also found that the UL9 protein in the dimer form can bind simultaneously to two DNA fragments, each containing specific binding sites for the protein. The interaction of the recombinant UL9 protein with the 63-mer double- and single-stranded oligonucleotides OriS and OriS*, which correspond to the origin of replication of herpes simplex virus, has been investigated. From the titrations of OriS and OriS* with ethidium bromide in the presence and absence of the UL9 protein, the equilibrium affinity constants of the protein binding to OriS and OriS* have been determined. A DNase I footprinting study showed that bis-netropsins exhibit preference for binding to the AT cluster in the origin of replication OriS and inhibit the fluctuation opening of AT base pairs in the AT cluster. The drugs also prevent formation of an intermediate conformation of OriS* that involves a disordered tail at the 3′ end and stable Box I-Box III hairpin to which the UL9 helicase selectively binds. The stabilization by bis-netropsins of the AT-rich hairpin at its 3′ end can inhibit the helicase activity. It was concluded that the antiviral activity of bis-netropsins may be associated with the inhibitory effects of bis-netropsins on these two stages of the reaction catalyzed by helicase UL9.