Host Proteins Involved in Integration and Excision
As mentioned above, the integration reaction requires supercoiled DNA or relaxed circular DNA plus ATP and Mg++. This suggests that the cell extracts contain a protein that can supercoil DNA using ATP for the required energy. Indeed, this proved to be so. By following the ability of an extract either to promote the integration reaction with relaxed circular substrates, or more simply, to produce supercoiled DNA from relaxed circular DNA, Gellert and coworkers identified and purified DNA gyrase. As explained, this enzyme introduces negative superhelical twists in covalently closed DNA circles.
Further study of the in vitro integration reaction revealed the require-ment for yet another host protein. This protein is directly involved in the integration and excision reactions. Extensive purification and char-acterization of this protein, which is called IHF, for integration host factor, showed it to be a dimer of rather small subunits, 11,000 and 9,500 molecular weight. The genes coding for these peptides, himA and himD, had been identified genetically from host mutations that block phage integration. It is surprising that cells should possess a nonessential protein that is required for integration of lambda. The IHF protein assists lambda phage integration and excision by helping the DNA bend into the complicated intasome structure that will be discussed below. This protein plays a similar role in other systems in which DNA bending is required.
Another host protein, called FIS, is also involved in the excision process. As the intracellular levels of this protein vary with the growth condition of the cells, it likely plays an important role in directing the phage into either the lytic or the lysogenic mode of existence, but it regulates only when Xis is present in limited amounts.