Single base-pair precision and structural rigidity in a small IHF-induced DNA loop.
The prokaryotic integration host factor (IHF) is a DNA-bending protein that binds to specific DNA sites as a heterodimer. Genetic and mutational analyses have previously identified asymmetric protein-DNA contacts by the individual subunits. By exploiting the unique sequence and positional context of one IHF binding site, H' in Lambda attachment sites (att sites), we have identified a symmetry element of binding and have localized the functional bend center to the center of this symmetry. A shift of the H' bend center by a single base-pair to the right or to the left within the very tight loop formed with Lambda integrase (Int) and IHF in att-site "intasomes" severely reduces recombination. This suggests that a precise, but wrongly positioned, DNA bend within a loop of constant length negatively influences the juxtaposition or "phasing" of the core-type and arm-type Int binding sites by differentially affecting the length of each leg of the loop. Furthermore, ten base-pair insertions within this loop that should not interfere with correct helical phasing are sensed in a position-dependent manner. Distal insertions abolish recombination, whereas proximal or double insertions (in both legs of the loop) are well tolerated.
Nunes-Düby
S E
SE
Division of Biology & Medicine, Brown University, Providence, RI 02912, USA.
Smith-Mungo
L I
LI
Landy
A
A
D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D001482:Base Composition; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D034982:Integration Host Factors; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D011995:Recombination, Genetic; D015183:Restriction Mapping
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D034982:Integration Host Factors; D004247:DNA