Premium
Effects of a High‐Affinity Antibody Fragment on DNA Polymerase Reactions Near a (6–4) Photoproduct Site
Author(s) -
Kobayashi Hiroyuki,
Sato Kousuke,
Komatsu Yasuo,
Morioka Hiroshi,
Stewart Jon D.,
Tsurimoto Toshiki,
Ohtsuka Eiko
Publication year - 1999
Publication title -
photochemistry and photobiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.818
H-Index - 131
eISSN - 1751-1097
pISSN - 0031-8655
DOI - 10.1111/j.1751-1097.1999.tb03278.x
Subject(s) - chemistry , fragment (logic) , polymerase , dna , microbiology and biotechnology , dna polymerase , antibody , stereochemistry , biophysics , biochemistry , biology , genetics , computer science , programming language
— Pyrimidine (6–4) pyrimidone photodimers are major photoproducts that have mutagenic and carcinogenic consequences. One major reason for these biological effects of (6–4) photoproducts may be base mispairing/DNA replication errors due to hydrogen bonding to bases opposite these damaged sites. We synthesized a modified 41‐mer DNA containing a (6–4) photoproduct using a preformed building block, then employed it as a template for primer extension reactions catalyzed by Klenow fragment and DNA polymerases α, β and δ (pol α, pol β and pol δ). None of these DNA polymerases were able to bypass the (6–4) photoproduct and elongation terminated at or near the 3′‐pyrimidone of the photoproduct, depending on the dNTP concentration. When a single‐chain Fv (scFv) with high affinity for the (6–4) photoproduct was included in the polymerization reaction, DNA synthesis was inhibited at base positions four, six, eight or eight nucleotides prior to the 3′‐pyrimidone by Klenow fragment, pol α, pol β or pol δ, respectively. These results suggest that the scFv can bind to the template DNA containing a (6–4) photoproduct and inhibit extension reactions by polymerases.