AFFINITY MODIFICATION OF HUMAN-IMMUNODEFICIENCY-VIRUS REVERSE-TRANSCRIPTASE AND DNA-TEMPLATE BY PHOTOREACTIVE DCTP ANALOGS

New base-substituted analogs of dCTP containing an azido group have be en synthesized and applied to a selective photoaffinity modification o f HIV-RT (p66/p51 heterodimer). The labeling of only the 66 kDa subuni t of HIV-RT was detected when the enzyme was first irradiated with the analogs and then template (5'-(d)GGTTAAATAAAATAGTAAGAATGTATAGCCCCTACC A-3') and 5' P-32 end-labeled 3'-(d)TTACATATCGGGGATGGT-5' primer were added. The 5' P-32 end-labeled primer elongated by dCTP analogs in the presence of both HIV-RT and DNA template is able to modify both subun its 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.