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Alternative conformations of human replication protein A are detected by crosslinks with primers carrying a photoreactive group at the 3′‐end
Author(s) -
Lavrik O.I.,
Kolpashchikov D.M.,
Nasheuer H.-P.,
Weisshart K.,
Favre A.
Publication year - 1998
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(98)01544-0
Subject(s) - primer (cosmetics) , primer extension , duplex (building) , dna polymerase , dna , dna replication , replication protein a , polymerase , protein subunit , base pair , nucleotide , rolling circle replication , chemistry , microbiology and biotechnology , biology , biochemistry , dna binding protein , base sequence , gene , organic chemistry , transcription factor
To analyze the influence of single‐stranded template extension of DNA duplex on the conformation of human replication protein A (RPA) bound to DNA we have designed two template‐primer systems differing by the size of the single‐stranded template tail (9 and 19 nucleotides (nt)). Base‐substituted photoreactive dUTP analogs were used as substrates for elongation of radiolabeled template‐primer by DNA polymerase β in the absence or in the presence of RPA. Following UV‐crosslinking it was demonstrated that the pattern of RPA subunit labeling and consequently RPA arrangement near the 3′‐end of the primer is stronlgly dependent upon the length of the template extension.