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- Title
A New Binary System for Photosensitized Labeling of DNA Polymerases in Nuclear Extract.
- Authors
Lebedeva, N. A.; Rechkunova, N. I.; Dezhurov, S. V.; Khodyreva, S. N.; Favre, A.; Lavrik, O. I.
- Abstract
A binary system of reagents was used for photosensitized labeling of proteins of bovine testis nuclear extract. A dUTP analog containing 4-azido-2,5-difluoro-3-chloropyridyl group (FAP-dUTP) was used for the first time as a component of the binary system, and a dUTP analog containing the pyrenyl group (Pyr-dUTP) was used as a photosensitizer. Photoaffinity labeling of proteins of nuclear extract was performed using the radioactively labeled DNA duplex with the photoreactive FAP group at the 3"-end of elongating DNA strand and analog of the deoxyribose phosphate residue (3-hydroxy-2-hydroxymethyltetrahydrofuran (F) 5"-phosphate) at the 5"-end of the nick. Such structure is formed by the action of nuclear extract enzymes from the initial DNA duplex containing a synthetic apurine/apyrimidine site and is a photoreactive analog of a long-patch base excision repair intermediate. UV-irradiation modified a limited number of proteins of the nuclear extract. As shown using specific antibodies, the new binary system of photoreagents increases the efficiency of DNA polymerase β labeling.
- Subjects
BINARY number system; CHEMICAL reagents; PROTEINS; TESTIS; PHOTOSENSITIZERS; DNA
- Publication
Biochemistry (00062979), 2003, Vol 68, Issue 4, p476
- ISSN
0006-2979
- Publication type
Article
- DOI
10.1023/A:1023616400794