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 mutati onal analyses have previously identified asymmetric protein-DNA contac ts by the individual subunits. By exploiting the unique sequence and p ositional context of one IHF binding site, H' in Lambda attachment sit es (att sites), we have identified a symmetry element of binding and h ave localized the functional bend center to the center of this symmetr y. 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 art-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 differe ntially 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. Dis tal insertions abolish recombination, whereas proximal or double inser tions (in both legs of the loop) are well tolerated. (C) 1995 Academic Press Limited