Acircular chromosome is a chromosome inbacteria,archaea,mitochondria, andchloroplasts, in the form of a molecule of circular DNA, unlike thelinear chromosome of mosteukaryotes.
Most prokaryote chromosomes contain a circular DNA molecule. This has the major advantage of having no free ends (telomeres) to theDNA. By contrast, most eukaryotes have linear DNA requiring elaborate mechanisms to maintain the stability of the telomeres andreplicate the DNA. However, a circular chromosome has the disadvantage that after replication, the two progeny circular chromosomes can remain interlinked or tangled, and they must be extricated so that each cell inherits one complete copy of the chromosome duringcell division.
The circular bacteria chromosome replication is best understood in the well-studied bacteriaEscherichia coli andBacillus subtilis. Chromosome replication proceeds in three major stages: initiation, elongation and termination. The initiation stage starts with the ordered assembly of "initiator" proteins at the origin region of the chromosome, calledoriC. These assembly stages are regulated to ensure that chromosome replication occurs only once in each cell cycle. During the elongation phase of replication, theenzymes that were assembled at oriC during initiation proceed along each arm (replichore) of the chromosome, in opposite directions away from the oriC, replicating the DNA to create two identical copies. This process is known as bidirectional replication. The entire assembly of molecules involved in DNA replication on each arm is called areplisome. At the forefront of the replisome is aDNA helicase that unwinds the two strands of DNA, creating a movingreplication fork. The two unwound single strands of DNA serve as templates forDNA polymerase, which moves with the helicase (together with other proteins) to synthesise a complementary copy of each strand. In this way, two identical copies of the original DNA are created. Eventually, the two replication forks moving around the circular chromosome meet in a specific zone of the chromosome, approximately opposite oriC, called the terminus region. The elongation enzymes then disassemble, and the two "daughter" chromosomes are resolved before cell division is completed.
TheE. coliorigin of replication, calledoriC consists ofDNA sequences that are recognised by theDnaA protein, which is highlyconserved amongst differentbacterial species. DnaA binding to the origin initiates the regulated recruitment of other enzymes andproteins that will eventually lead to the establishment of two complete replisomes for bidirectional replication.[1]
DNA sequence elements withinoriC that are important for its function include DnaA boxes, a 9-mer repeat with a highly conserved consensus sequence 5' – TTATCCACA – 3',[2] that are recognized by the DnaA protein. DnaA protein plays a crucial role in the initiation of chromosomal DNA replication.[3] Bound to ATP, and with the assistance of bacterialhistone-like proteins [HU] DnaA then unwinds an AT-rich region near the left boundary oforiC, which carries three 13-mer motifs,[4] and opens up thedouble-stranded DNA for entrance of other replication proteins.[5]
This region also contains four "GATC"DNA unwinding element sequences that are recognized byDNA adenine methylase (Dam), an enzyme that modifies the adenine base when this sequence is unmethylated or hemimethylated. Themethylation ofadenines is important as it alters the conformation of DNA to promote strand separation,[6] and it appears that this region oforiC has a natural tendency to unwind.[7]
DnaA then recruits the replicativehelicase,DnaB, from the DnaB-DnaC complex to the unwound region to form the pre-priming complex.[8] After DnaB translocates to the apex of each replication fork, the helicase both unwinds the parental DNA and interacts momentarily withprimase.[9]
In order for DNA replication to continue, single strandedbinding proteins are needed to prevent the single strands of DNA from forming secondary structures and to prevent them fromre-annealing. In addition,DNA gyrase is needed to relieve the topological stress created by the action of DnaB helicase.
When the replication fork moves around the circle, a structure shaped like the Greek lettertheta Ө is formed.John Cairns demonstrated thetheta structure of E. coli chromosomal replication in 1963, using an innovative method to visualize DNA replication. In his experiment, heradioactively labeled the chromosome by growing his cultures in a medium containing 3H-thymidine. Thenucleoside base was incorporated uniformly into the bacterial chromosome. He then isolated the chromosomes by lysing the cells gently and placed them on anelectron micrograph (EM) grid which he exposed toX-ray film for two months. This Experiment clearly demonstrates the theta replication model of circular bacterial chromosomes.[10]
As described above, bacterial chromosomal replication occurs in a bidirectional manner. This was first demonstrated by specifically labelling replicating bacterial chromosomes withradioactive isotopes. The regions of DNA undergoing replication during the experiment were then visualized by usingautoradiography and examining the developed film microscopically. This allowed the researchers to see where replication was taking place. The first conclusive observations of bidirectional replication was from studies of B. subtilis.[11] Shortly after, the E. coli chromosome was also shown to replicate bidirectionally.[12]
TheE. coliDNA polymerase III holoenzyme is a 900 kD complex, possessing an essentially adimeric structure. Eachmonomeric unit has a catalytic core, adimerization subunit, and aprocessivity component .[13] DNA Pol III uses one set of its core subunits to synthesize theleading strand continuously, while the other set of core subunits cycles from oneOkazaki fragment to the next on the loopedlagging strand. Leading strand synthesis begins with the synthesis of a shortRNA primer at the replication origin by the enzyme Primase (DnaG protein).
Deoxynucleotides are then added to this primer by a single DNA polymerase III dimer, in an integrated complex with DnaB helicase. Leading strand synthesis then proceeds continuously, while the DNA is concurrently unwound at the replication fork. In contrast, lagging strand synthesis is accomplished in short Okazaki fragments. First, an RNA primer is synthesized by primase, and, like that in leading strand synthesis, DNA Pol III binds to the RNA primer and addsdeoxyribonucleotides.
When the synthesis of an Okazaki fragment has been completed, replication halts and the core subunits of DNA Pol III dissociates from the β sliding clamp [B sliding clap is the processivity subunit of DNA Pol III].[14] The RNA primer is removed and replaced with DNA byDNA polymerase I [which also possesses proofreadingexonuclease activity] and the remaining nick is sealed byDNA ligase, which then ligates these fragments to form the lagging strand.
A substantial proportion (10-15%) of the replication forks originating at oriC encounter aDNA damage or strand break when cells are grown under normal laboratory conditions (without an exogenous DNA damaging treatment).[15] The encountered DNA damages are ordinarily processed byrecombinational repair enzymes to allow continued replication fork progression.[15]
Termination is the process of fusion of replication forks and disassembly of the replisomes to yield two separate and completeDNA molecules. It occurs in the terminus region, approximately opposite oriC on the chromosome (Fig 5). The terminus region contains several DNA replication terminator sites, or "Ter" sites. A special "replication terminator" protein must be bound at the Ter site for it to pause replication. Each Ter site has polarity of action, that is, it will arrest a replication fork approaching the Ter site from one direction, but will allow unimpeded fork movement through the Ter site from the other direction. The arrangement of the Ter sites forms two opposed groups that forces the two forks to meet each other within the region they span. This arrangement is called the "replication fork trap."[16]
The Ter sites specifically interact with the replication terminator protein calledTus inE. coli.[17] The Tus-Ter complex impedes theDNA unwinding activity of DnaB in an orientation-dependent manner.[18]
Replication of the DNA separating the opposing replication forks leaves the completed chromosomes joined as 'catenanes' or topologically interlinked circles. The circles are not covalently but mechanically linked, because they are interwound[check spelling] and each is covalently closed. The catenated circles require the action oftopoisomerases to separate the circles (decatenation). InE. coli, DNA topoisomerase IV plays the major role in the separation of the catenated chromosomes, transiently breaking both DNA strands of one chromosome and allowing the other chromosome to pass through the break.
There has been some confusion about the roleDNA gyrase plays in decatenation. To define the nomenclature, there are two types of topoisomerases: type I produces transient single-strand breaks in DNA and types II produces transient double-strand breaks. As a result, the type I enzyme removessupercoils from DNA one at a time, whereas the type II enzyme removes supercoils two at a time. The topo I of bothprokaryotes and eukaryotes are the type I topoisomerase. The eukaryotic topo II, bacterial gyrase, and bacterial topo IV belong to the type II.
DNA gyrase also has topoisomerase type II activity; thus, with it being a homologue of topoisomerase IV (also having topoisomerase II activity) we expect similarity in the two proteins' functions. DNA gyrase's preliminary role is to introduce negative super coils into DNA, thereby relaxing positive supercoils that form during DNA replication. Topoisomerase IV also relaxes positive supercoils, therefore, DNA Gyrase and topoisomerase IV play an almost identical role in removing the positive supercoils ahead of a translocating DNA polymerase, allowing DNA replication to continue unhindered by topological strain.[19]
DNA gyrase is not the sole enzyme responsible for decatenation. In an experiment byZechiedrich, Khodursky and Cozzarelli in 1997, it was found that topoisomerase IV is the only important decatenase of DNA replication intermediates in bacteria.[20] When DNA gyrase alone was inhibited, most of the catenanes were unlinked. However, when Topoisomerase IV alone was inhibited, decatenation was almost completely blocked. This suggests that Topoisomerase IV is the primary protein for decatenation of interlinked chromosomesin vivo, with DNA gyrase playing a minor role.
Several groups of bacteria, includingBrucella,Paracoccus denitrificans, andVibrio have multiple circular chromosomes.
This is based on an article by Imalda Devaparanam and David Tribe made available under CC by SA licensing conditions from a university course activity at the Department of Microbiology and Immunology, University of Melbourne, 2007.[citation needed]This article incorporates material from theCitizendium article "Replication of a circular bacterial chromosome", which is licensed under theCreative Commons Attribution-ShareAlike 3.0 Unported License but not under theGFDL.
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