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Nearly all DNA polymerases require processivity factors to ensure continuous incorporation of nucleotides. Processivity factors are specific for their cognate DNA polymerases. For this reason, the vaccinia DNA polymerase (E9) and the proteins associated with processivity (A20 and D4) are excellent therapeutic targets. In this study, we show the utility of stepwise rapid plate assays that i) screen for compounds that block vaccinia DNA synthesis, ii) eliminate trivial inhibitors, e.g. DNA intercalators, and iii) distinguish whether inhibitors are specific for blocking DNA polymerase activity or processivity. The sequential plate screening of 2,222 compounds from the NCI Diversity Set library yielded a DNA polymerase inhibitor (NSC 55636) and a processivity inhibitor (NSC 123526) that were capable of reducing vaccinia viral plaques with minimal cellular cytotoxicity. These compounds are predicted to block cellular infection by the smallpox virus, variola, based on the very high sequence identity between A20, D4 and E9 of vaccinia and the corresponding proteins of variola.
© <2008>. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/
Poxvirus, vaccinia virus, DNA polymerase, processivity factor, antiviral inhibitors, high throughput screening, rapid plate assay, smallpox
Silverman, J. E., Ciustea, M., Shudofsky, A. M., Bender, F., Shoemaker, R. H., & Ricciardi, R. P. (2008). Identification of Polymerase and Processivity Inhibitors of Vaccinia DNA Synthesis Using a Stepwise Screening Approach. Antiviral Research, 80 (2), 114-123. http://dx.doi.org/10.1016/j.antiviral.2008.05.010
Date Posted: 24 February 2022
This document has been peer reviewed.