Restriction-Modification Systems
|
|
|
|
Control of restriction vs. modification:Type I restriction-modification enzymes are multifunctional, multisubunit enzymes that provide their host bacteria with protection against invading DNA. This protection is accomplished through a very efficient cleavage of foreign DNA using a complex mechanism involving hydrolysis of ATP and subsequent translocation of the substrate DNA. In addition, the same enzyme provides protection against cleavage for the host chromosome by methylating the target recognition sites. However, the restriction (cleavage) activity of the enzyme must be carefully controlled both in terms of what constitutes substrate DNA ("foreign" versus "host") and through temporal control of the two activities when the R-M system is transferred to a new host (modification must precede restriction to allow establishment of the R-M system). The mechanism by which one of these enzymes differentiates between "foreign" and "host" DNA is described by Firman et al. (2000), who also discuss recent evidence showing post-translational control as the primary mechanism for temporal control of restriction. The importance of subunit assembly in these processes is detailed and extended to include novel assemblies with unexpected function.
Temporal control of R-M enzymes is required following transfer of the R-M system between different strains of bacteria (e.g. via conjugation). It has been clearly demonstrated (Prakash-Cheng & Ryu Junichi 1993) that modification activity (DNA methylation) always precedes restriction activity (DNA cleavage) following conjugation of the R-M system into a new host bacteria. Two systems have been studied in an attempt to understand how this control is produced. The Type IA enzyme EcoKI is controlled through proteolytic activity against the HsdR subunit of the active, fully assembled restriction endonuclease (Makovets et al., 1998, 1999). The Type IC enzyme EcoR124I is controlled through subunit assembly, with the final step in the assembly pathway being formation of a weak R2-complex. This requires "build-up" of the concentration of the HsdR subunit, within the cell following conjugal transfer, before the R2-complex can assemble and produce restriction activity. During this period of time (of synthesis of HsdR) methylation of the host DNA can occur. In addition, it seems likely that Type I R-M enzymes also use cellular localisation to separate the functionality of the multifunctional enzyme. This has been shown in studies by Holubova et al., who have demonstrated family differences in the cellular localisation of these enzymes. Finally, there is a continual "battle" between the ability of bacteriophage or plasmids to infect a recipient cell and the cell's own requirement for survival. As already mentioned R-M systems present a first barrier to infection by "foreign" DNA and are the first line of defence in this battle, but both bacteriophage and plasmids have evolved anti-restriction systems to overcome these barriers. |
|
Send feedback about the site including suggestions for content via
Email.
|
The
ability to differentiate
between "host" and "foreign" DNA is dependent upon the methylation status of the
DNA at the recognition sequence. The preferred substrate for