Читать книгу Snyder and Champness Molecular Genetics of Bacteria - Tina M. Henkin - Страница 83

After replication of the chromosome initiates in the oriC region and proceeds around the circular chromosome in both directions, the two replication forks must meet somewhere on the chromosome and the two daughter chromosomes must separate. In some bacteria, including E. coli and B. subtilis, a specific system exists to control the region where replication forks meet. What happens in other organisms is less clear. As with most cellular processes, the process of termination of chromosome replication is especially well understood in E. coli. In this bacterium, termination is facilitated by DNA sequences, called ter sites, that are only ~22 bp long. These sites act somewhat like the one-way gates in an automobile parking lot, allowing the replication forks to pass through in one direction but not in the other.

Figure 1.15 Initiation of replication at the Escherichia coli origin (oriC) region. DnaA is always bound to three DnaA boxes within oriC, even when DnaA is in its non-ATP-bound state acting as an origin recognition complex. About a dozen DnaA-ATP proteins bind to the origin, possibly by forming a type of helical filament and opening the helix and the DNA-unwinding element. DnaC helps the DnaB helicase to bind. The DnaG primase synthesizes RNA primers, initiating replication.

Figure 1.16 shows how the one-way nature of ter sequences can cause replication to terminate in a specific region of the chromosome. In the illustration, two ter sites called terA and terB bracket the termination region. Replication forks are unaffected by the terA site in the clockwise direction but are terminated in the counterclockwise direction. The opposite is true for terB. Thus, the clockwise-moving replication fork progresses through terA, but if it gets to terB before it meets the counterclockwise-moving fork, it stalls, because it cannot move clockwise through terB. Similarly, the replication fork moving in the counterclockwise direction stalls at the terA site and waits for the clockwise-moving replication fork. When the counterclockwise and clockwise replication forks meet, at terA, terB, or somewhere between them, the two forks terminate replication, releasing the two daughter DNAs. In E. coli, it is known that most DNA replication termination occurs at one ter site, possibly because it is oriented to terminate replication forks traveling in the clockwise direction, which is shorter in most laboratory E. coli strains.

Figure 1.16 Termination of chromosome replication in Escherichia coli. (A) The replication forks that start at the origin of chromosomal replication (oriC) can traverse terA and terB in only one direction, opposite that indicated by the black arrows. (B) When they meet, between or at one of the two clusters, chromosome replication terminates. f_L is the fork that initiated to the left and moved in a counterclockwise direction. f_R is the fork that initiated to the right and moved in a clockwise direction. Adapted from Camara JE, Crooke E, in Higgins MP, ed, The Bacterial Chromosome (ASM Press, Washington, DC, 2005), with permission.

Encountering a ter DNA sequence, by itself, is not sufficient to stop the replication fork. A protein is also required to terminate replication at ter sites. The protein that works with ter sites, called the terminus utilization substance (Tus) in E. coli and the replication terminator protein (RTP) in B. subtilis, binds to the ter sites and stops the replicating helicase (DnaB in E. coli) that is separating the strands of DNA ahead of the replication fork. In both E. coli and B. subtilis, multiple ter sites bracket the terminus region, helping to ensure that replication proceeds from oriC to the terminus region (see Box 1.1). While the ter systems are not absolutely essential for E. coli and B. subtilis growing in the laboratory setting, they are important for other aspects of genome stability that are important for maintaining genome integrity over time in the natural environment (see Rudolph et al., Suggested Reading). It has been argued that the active termination systems involving ter sites and a trans acting protein originated in plasmids (chapter 5) and were domesticated in some branches of bacteria for use in the bacterial chromosome (see Galli et al., Suggested Reading).