P1 PLASMID PARTITION - BINDING OF P1 PARB PROTEIN AND ESCHERICHIA-COLI INTEGRATION HOST FACTOR TO ALTERED PARS SITES

The Escherichia coli integration host factor (IHF) participates in P1 plasmid partition by assisting the interaction of P1 ParB protein with its specific site, parS. Together they form an extremely high-affinit y protein-DNA complex, in which parS DNA is wrapped around a core of P arB and IHF protein in a precise three-dimensional conformation. We ha ve investigated the interaction of ParB and IHF with mutant DNA sites, to examine protein specificity and cooperativity. The results indicat e that ParB specifically recognizes two separate types of sequence rep eats in its minimal binding site in one half of the parS site. The aff inity of ParB or IHF for parS is much greater in the presence of the o ther protein. Mutations that decrease ParB or IHF binding to parS have relatively minor defects in vivo, because each protein still binds we ll to parS in the presence of the other protein. We observed that ParB acts better when provided in cis than in trans to parS in vivo. Our e xperiments suggest that in vivo, the local concentration of ParB prote in near the plasmid is high, so that ParB can act reasonably well to p romote partition in cells without IHF. However, this activity is lower than in wild-type cells, indicating that IHF is essential for long-te rm plasmid stability.