Growth fork

Schematic diagrams of two different modes of DNA synthesis at the growth fork(s). In unidirectional replication (a) one growth fork occurs in bidirectional replication (b) two occur. Red indicates regions containing newly synthesized DNA.

At the growth fork it is necessary that the parental double helix be unwound to present further single-stranded regions to serve as templates for continued replication. The dnaB protein of E. coli is believed to be a helicase that directly catalyzes this process. 

Single-strand binding protein (SSB) is found in abundance in E. coli, and it is believed to be bound in mass at the replication fork. This fact and other evidence described later on indicate that it plays an important role at the growth fork. 

Two aspects of E. coli chromosomal replication still to be considered are initiation and termination. From what has been said we conjecture that replication initiates at a unique site, proceeds bidirectionally, and terminates at a point where the two oppositely advancing growth forks meet. To study initiation it was first necessary to isolate that segment... 

Growth fork. The region on a DNA duplex molecule where synthesis is taking place. It resembles a fork in shape because it consists of a region of duplex DNA connected to a region of unwound single strands. 

Replication fork. The Y-shaped region of DNA at the site of DNA synthesis also called a growth fork. 

Figure 11.6 Multiple origin of replication forks in eukaryotic DNA replication. Termination of replication occurs where two growth forks come together.

DNA synthesis occurs in both directions at each of the rep-licating forks. Once a DNA strand has been primed, synthesis toward the replicating fork can be visualized as continuous. Growth of the opposite,... 

Clifton, K.H. and Sridharan, B.N. (1975). Endocrine factors and growth, page 249 in Cancer A Comprehensive TYeatise, Vol 3, BECKER, EE, Ed. (Plenum Press, New "fork). 

The circular chromosome of an E. coli cell contains 4.6 x 106 base pairs. If a replication fork moves at a rate of 1000 nucleotides per second, how much time will be required for replication of the DNA Cells of E. coli can divide every 20 minutes under favorable conditions. How can you explain this rapid rate of growth ... 

Growth during Replication Is Bidirectional Growth at the Replication Forks Is Discontinuous Proteins Involved in DNA Replication Characterization of DNA Polymerase I in Vitro Crystallography Combined with Genetics to Produce a Detailed Picture of DNA Poll Function... 

Models for synthesis at the replication fork, (a) Continuous synthesis on both strands. Note that both growth arrows are pointing in the same direction, which would require growth in the 5 — 3 direction on one strand and in the 3 — 5 direction on the other. If growth occurs only in the 5 — 3 direction, synthesis would have to be discontinuous on one strand, as in (b). Alternatively, it could be discontinuous on both strands (c). 

Fig. 16-14 Events involved in DNA chain growth at the replication fork.

Temperature-sensitive mutations, in particular, have been very valuable in helping to define many of the proteins involved in replication. Several of these proteins have already been discussed. Temperature-sensitive mutations take effect at a certain temperature, e.g., 42-47°C, and not at another, e.g., 30°C or less. Mutations that affect replication are called dna mutations. Many that have been identified in E. coli code for various proteins associated with DNA chain growth at the replication fork. For example, the gene dnaG codes for primase (the DnaG protein) which has already been discussed. Some, however, code for proteins involved also or exclusively with the initiation of a cycle of replication at oriC. Examples of these are dnaA, B and C. 

Many of the proteins and enzymes involved in initiation at replication origins and DNA chain growth at replication forks have the same biochemical activities as their counterparts in bacteria. However, the situation regarding terminators and terminator proteins is less clear. Whether they exist to delineate to any extent individual replicons or clusters of replicons is not known. In the case of eukaryotic chromosomes, however, there is a special mechanism to replicate their ends which are known as telomeres. 

Question What is known about the process of DNA chain growth at replication forks in eukaryotes ... 

The structure of replication forks in eukaryotes is essentially the same as in bacteria. Chain growth is continuous on the leading strand and discontinuous on the lagging strand. There are equivalents of the polymerases, helicase, primase, SSB, etc., but there are clearly some differences. For example, two different polymerases, DNA polymerase 8 and DNA polymerase a, function on the leading and lagging strand, respectively. Also, the mitochondrion has its own DNA polymerase. 

A number of nucleotide analog compounds function by blocking further chain growth at the replication fork. The 2 3 -dideoxynucleosides can be converted to the triphosphates (ddNTPs). In the case of bacteria, these are incorporated onto the 3 -hydroxyl end of a growing DNA chain, and because the new end now lacks a 3 hydroxyl, no further additions can occur. They are used in conjunction with DNA polymerase I in the dideoxy method of Sanger for determining DNA sequences. 

A multiforked bacterial chromosome is one that contains more than two replication forks and results from reinitiation at the daughter origins within a replication bubble. In this situation, cycles of replication are completed at more frequent intervals, and this gives shorter generation times. It occurs under conditions of fast growth, induced by nutritionally rich media. 

DNA chain growth occurs on both daughter arms at a replication fork. On one arm, chain growth occurs continuously (5 —>3 ), in the same direction as fork movement. On the other arm, chain growth occurs in separate short pieces (5 — 3 ) and in the direction opposite to fork movement. The short pieces (nascent or Okazaki fragments) subsequently join. Replication in the latter fashion is known as discontinuous DNA replication. 

Unlike the DNA polymerases, RNA polymerase is able to initiate a new RNA chain, using DNA as a template (Chap. 17). The DNA polymerases are able to extend the DNA from an RNA primer. In discontinuous DNA chain growth, a particular type of RNA polymerase, called primase in E. coli, lays down short RNA primers at fairly regular base intervals, as unwinding of the helix at the replication fork proceeds. These primers are involved in the initiation of synthesis of nascent DNA chains by DNA polymerase. 

Samson, G., Herbert, S.K., Fork, D.C. and Laudenbach, D.E. 1994. Acclimation of the photosynthetic apparatus to growth irradiance in the mutant strain of Synechococcus lacking iron superoxide dismutase. Plant Physiol. 105, 287-294. 

Oxidative damage that leads to replication fork arrest is common during bacterial growth and probably occurs at least once during each cell division even under the most favorable conditions (48, 51). Most lesions are not dealt with by mutageiuc polymerases, but instead they are repaired by recombination, which is an error-free process. Recombinational repair can take many different paths and employs several different proteins, but ultimately, it involves high-fidelity DNA polymerases such as Pol I, Pol II, and/or Pol III (6, 49, 56). For additional information, we refer the reader to more thorough reviews of the various recombination repair pathways (44, 48-50,70). 

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