Cells and viruses use conservative site-specific recombination for a wide variety of biological functions. Some of these functions are discussed in the following sections. Many phage insert their DNA into the host chromosome during infection using this recombination mechanism. In other cases, site-specific recombination is used to alter gene expression. For example, inversion of a DNA segment can allow two alternative genes to be expressed. Site-specific recombination is also widely used to help maintain the structural integrity of circular DNA molecules during cycles of DNA replication, homologous recombination, and cell division.
A comparison of site-specific recombination systems reveais some general themes. All reactions depend critically on the assembly of the recombinase protein on the DNA, and the bringing together of the two recombination sites. For some recombination reactions this assembly is very simple, requiring only the recombinase and its DNA recognition sequences as just described for Cre. In contrast, other reactions require accessory proteins. These accessory proteins include so-called architectural proteins thai bind specific DNA sequences and bend the DNA. They organize DNA into a specific shape and thereby stimulate the recombination. Architectural proteins can also control the direction of a recombination reaction, for example, to ensure that integration of a DNA segment occurs while preventing the reverse reaction—DNA excision, Clearly, this type of regulation is essential for a logical biological outcome. Finally, we will also see thai recombinase» can be regulated by other proteins to control when a particular DNA rearrangement takes place and coordinate it with other cellular events.
Biological Roies of Site-Specific Recombination 30H
X Integrase Promotes the Integration and Excision of a Viral Genome into the Host Cell Chromosome
When bacteriophage X infects a host bacterium, a series of regulatory events result either in establishment of the quiescent lysogenic state or in phage multiplication, a process called lytic growth (see Chapters 16 and 21). Establishment of a lysogen requires the integration of the phage DNA into the host chromosome. Likewise, when the phage loaves the lysogenin state to roplicate and make new phage particles. 31 must excise its UNA from the host chromosome, The analysis of this integration/excision reaction provided the first molecular insights into site-specific recombination.
To integrate, the K integrase protein (Xint) catalyzes recombination between two specific sites, known as the att, or attachment, sites. The offPsite is on She phage DNA (P for phage) and the afîB site is in the. bacterial chromosome (0 for bacteria; see Figure 11-2). Xint is a tyrosine recombinase, and the mechanism of strand exchange follows the pathway described above for the Cre protein. Unlike Cre recombination, however, X integration requires accessory proteins to help the required protem-DNA complex to assemble. These proteins control the reaction to ensure that DNA integration and DNA excision occur at the right time in the phage life cycle. Wc will first consider the integration pathway find then look at how excision is triggered,
Important to the regulation of K integration is the highly asymmetric organization of the attP and attB sites (Figure 11-9). Both sites carry
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