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.
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