A screen in Escherichia coli for nucleoside analogs that target human immunodeficiency virus (HIV) reverse transcriptase: coexpression of HIV reverse transcriptase and herpes simplex virus thymidine kinase.
Human immunodeficiency virus (HIV) reverse transcriptase substitutes for temperature-sensitive DNA polymerase I (Pol Its) in Escherichia coli, providing a screen for anti-HIV reverse transcriptase nucleoside analogs in bacteria. Since phosphorylation of nucleosides in E. coli is limited to thymidine and its derivatives, we coexpressed herpes simplex virus thymidine kinase, an enzyme that phosphorylates a wide variety of nucleoside analogs, together with HIV reverse transcriptase. Coexpression of herpes simplex virus thymidine kinase and HIV reverse transcriptase rendered Pol Its cells sensitive to dideoxycytidine. Studies with different nucleoside analogs indicate that this bacterial screening system is able to select and identify nucleoside analogs that specifically target HIV reverse transcriptase.
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Joseph Gottstein Memorial Cancer Research Laboratory, Department of Pathology and Biochemistry, University of Washington, Seattle 98195, USA.
Loeb
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D000998:Antiviral Agents / Q000494:pharmacology*; D003001:Cloning, Molecular; D003561:Cytarabine / Q000494:pharmacology*; D004256:DNA Polymerase I; D004353:Drug Evaluation, Preclinical / Q000379:methods; D004926:Escherichia coli / Q000187:drug effects* / Q000201:enzymology; D015870:Gene Expression; D006678:HIV / Q000201:enzymology*; D007700:Kinetics; D010766:Phosphorylation; D012194:RNA-Directed DNA Polymerase / Q000096:biosynthesis / Q000378:metabolism; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism; D018894:Reverse Transcriptase Inhibitors; D018139:Simplexvirus / Q000201:enzymology*; D013937:Thymidine Kinase / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000378:metabolism; D016047:Zalcitabine / Q000494:pharmacology*; D015215:Zidovudine / Q000494:pharmacology*
D000998:Antiviral Agents; D011994:Recombinant Proteins; D018894:Reverse Transcriptase Inhibitors; D003561:Cytarabine; D015215:Zidovudine; D016047:Zalcitabine; D013937:Thymidine Kinase; D012194:RNA-Directed DNA Polymerase; D004256:DNA Polymerase I