site-specific recombination and replicon stability

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In this research program, we wish to examine how a bacteria's genetic material separates before the cell divides. In order for this to happen, the cell must ensure that its DNA has completely replicated and has been equally partitioned into each daughter cell before the division process terminates. A problem often arises during this process is the recombination of the 2 daughter chromosomes prior to cell division, leading to the formation of a chromosome dimer. If this double chromosome cannot be effectively separated, the daughter cells will not receive identical copies of the bacterial genome, leading to cell death. To overcome this problem, bacteria have devised a way to reconvert double chromosomes into 2 single, independant, identical copies using a genetic process called site-specific recombination. This process ensures tha tthis recombination event occurs at the right time (prior to cell division) and at the right place (at the cell septum, the junction point between the newly formed daughter cells). In the majority of bacteria, this process is performed by 2 cellular proteins, XerC and XerD, which act in concert at a specific chromosomal site called dif. Recently, it was shown that certain bacteria like Streptococci, Lactococci, Helicobacter and Campylobacter use a single Xer protein to perform this identical process. We are interested in how the 'double' XerCD system, and the 'single' Xer system (called XerS/H) work at the molecular level (how the proteins bind DNA, what bases are critical, and how the recombination reaction proceeds after DNA binding). These single and double recombinase systems seem to have evolved divergently, and we have recently found that certain bacteria possess both single and double recombinase systems. We are interested in determining how these systems have evolved, and, in the case of bacteria which possess both systems, finding out which one is essential. The main thrust of our work is to fully understand how this important cellular process works at the molecular level, and how it is regulated both temporally and spatially.