Site-Specific Recombination

Alignment of sites for homologous recombination occurs via DNA - DNA (base-pairing) interactions. Another important class of recombination reactions is called site-specific recombination. It is directed by highly specific DNA - protein interactions, although a short stretch of DNA homology occurs at the actual site of cutting and resealing. Information about site-specific recombination is most advanced for the mechanism by which phages, such as , become integrated at specific sites on an infected bacterium's chromosome.

The chromosome integrates at a specific site on the E. coli chromosome, attB, which maps between genes involved in galactose utilization and biotin synthesis (the gal and bio markers), as illustrated in Figure 25.17. Integration occurs at a specific site on the phage chromosome called attP.

Two proteins are required for this site-specific recombination. They are as follows:

> 1. Phage integrase (Int) - the product of the int gene; and

> 2. Integration Host Factor (IHF)--an E. coli protein.

Phage DNA must be supercoiled for the recombination to occur. Supercoiling, plus distortion created by Ihf binding to specific sites in attP, facilitates Int binding at adjacent sites. The nucleoprotein structure is called an intasome, which aligns with attB (also bound with Int). At the core of the and E. coli sequences is a 15-base region of complete homology (Figure 25.31). In each of these sequences, Int creates a staggered cut, with a 7-nucleotide overlap. The ends then exchange to form a Holliday junction and a DNA ligase activity of Int joins the ends covalently.

Upon integration, is dormant. Later, changes in the cell activate the virus which excises from the host genome by a reversal of the above steps to yield a circular virus. In this reaction, a protein called Xis is required in addition to Int and IHF.

See also: Recombination, Homologous Recombination

INTERNET LINK Recombination Tutorial