Abstract
New base-substituted analogs of dCTP containing an azido group have been synthesized and applied to a selective photoaffinity modification of HIV-RT (p66/p51 heterodimer). The labeling of only the 66 kDa subunit of HIV-RT was detected when the enzyme was first irradiated with the analogs and then template (5'-(d)GGTTAAATAAAATAGTAAGAATGTATAGCCCCTACCA-3') and 5' 32P end-labeled 3'-(d)TTACATATCGGGGATGGT-5' primer were added. The 5' 32P end-labeled primer elongated by dCTP analogs in the presence of both HIV-RT and DNA template is able to modify both subunits of HIV-RT and DNA template. This way of specific cross-linking to both DNA (RNA) template and HIV-RT opens up new possibilities to study the HIV-RT active site.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Affinity Labels*
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Base Sequence
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Binding Sites
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DNA / chemistry*
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DNA / metabolism
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Deoxycytosine Nucleotides / chemistry
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Deoxycytosine Nucleotides / metabolism
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Deoxycytosine Nucleotides / pharmacology*
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HIV / enzymology*
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Molecular Sequence Data
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Molecular Structure
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Photochemistry
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RNA-Directed DNA Polymerase / chemistry*
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RNA-Directed DNA Polymerase / metabolism
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Templates, Genetic
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Ultraviolet Rays
Substances
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5-(3-(4-azidotetrafluorobenzamido)allyl)-2'-deoxycytidine-5'-triphosphate
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Affinity Labels
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Deoxycytosine Nucleotides
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exo-N-(2-(4-azidotetrafluorobenzamido)ethyl)-deoxycytidine-5'-triphosphate
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2'-deoxycytidine 5'-triphosphate
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DNA
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RNA-Directed DNA Polymerase