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Potent DNA Strand Annealing Mediated by the T7 Single‐Stranded DNA Binding Protein gp2.5
Author(s) -
Hernandez Alfredo J.,
Richardson Charles C.
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.591.6
Subject(s) - dna , replication protein a , homologous recombination , dna clamp , branch migration , dna replication , oligonucleotide , chemistry , in vitro recombination , biology , biophysics , microbiology and biotechnology , gene , genetics , dna binding protein , polymerase chain reaction , complementary dna , holliday junction , reverse transcriptase , molecular cloning , transcription factor
Single‐stranded DNA binding proteins are ubiquitous in all domains of life and play essential roles in DNA replication, recombination, and repair. Bacteriophage T7 encodes a single‐stranded DNA binding protein, gp2.5, which is crucial for coordinating the synthesis of leading and lagging DNA strands. In addition to its role in DNA replication, T7 gp2.5 displays a potent homologous DNA strand annealing activity, which is necessary for genetic recombination. Gp2.5 enhances the annealing of DNA strands orders of magnitude over the spontaneous rate, and is considerably more efficient compared to other recombination proteins. The annealing of complimentary strands by gp2.5 is strictly specific for DNA. While the annealing reaction is somewhat tolerant to internal mismatches, it is very permissive to internal, unpaired loops. Here we provide a model for the mechanism of gp2.5‐mediated homologous strand annealing and explore its applicability in oligonucleotide‐mediated gene modification and the formation of higher‐order DNA structures in vitro .