Replication in prokaryotes

The overall process of DNA replication in prokaryotes and eukaryotes is compared in Figure 1-2-1. 

The mechanism of replication in eukaryotes is believed to be very similar to this. However, the details have not yet been completely worked out. The steps and proteins involved in DNA replication in prokaryotes are compared with those used in eukaryotes in Table 1-2-2. 

Although replication in prokaryotes is now well understood, many details in eukaryotes are still unclear. However, it is certain that the process is in principle similar. A simplified scheme of replication in the bacterium Escherichia coli is shown here. 

The process of eukaryotic DNA replication closely follows that of prokaryotic DNA synthesis. Some differences, such as the multiple origins of replication in eukaryotic cells versus single origins of replication in prokaryotes, have already been discussed. Eukaryotic single-stranded DNA-binding proteins and ATP-dependent DNA helicases have been identified, whose functions are analogous to those of the prokaryotic enzymes previously discussed. In contrast, RNA primers are removed by RNase H. 

Postreplicational mismatch repair has been found to correct errors in base substitution occurring during DNA replication in prokaryotes.48 This lowers the error rate for the polymerase from 1 in 106 to 107 to the observed range of values of 1 in 108 to 1010 in E. coli. How does the repair system know in this case which base in a mispair is the incorrect one The answer appears to be that the parent strand is tagged by methylation. A small proportion, some 0.2%, of the cytosine residues are methylated at the 5 position, and a similar proportion of the adenine residues are methylated at the 6 position. As methylation is a postreplicative event, the daughter strand is temporarily undermethylated after replication. 

The flow of genetic information involves biosyntheses of DNA, RNA, and proteins, known as replication, transcription, and translation, respectively. DNA replication in prokaryotes (Nossal, 1983) and eukaryotes (Campbell, 1985) are very similar, though eukaryotic replication is more complex (DePamphilis, 1996). A... 

Replication in eukaryotes is mechanistically similar to replication in prokaryotes but is more challenging for a number of reasons. One of them is sheer size E. coli must replicate 4.8 million base pairs, whereas a human diploid cell must replicate 6 billion base pairs. Second, the genetic information for . coli is contained on 1 chromosome, whereas, in human beings, 23 pairs of chromosomes must be replicated. Finally, whereas the E. coli chromosome is circular, human chromosomes are linear. Unless countermeasures are taken (Section 27.4.7), linear chromosomes are subject to shortening with each round of replication. 

Fig 11.14 Single bubble for DNA Replication in Prokaryotic Chromosomes Requirements for DNA Synthesis... 

DNA SYNTHESIS IN EUKARYOTES Although the principles of DNA replication in prokaryotes and eukaryotes have a great deal in common (e.g., semiconservative replication and bidirectional replicons), they also have significant differences. Not surprisingly, these differences appear to be related to the size and complexity of eukaryotic genomes. 

Another important difference between DNA replication in prokaryotes and in eukaryotes is that prokaryotic DNA is not complexed to histones, as is eukaryotic DNA. Histone biosynthesis occurs at the same time and at the same rate as DNA biosynthesis. In eukaryotic replication, histones are associated with DNA as it is formed. An important aspect of DNA replication in eukaryotes, specifically affecting humans, is described in the Biochemical Connections box on pages 282 and 283. 

Replication in eukaryotes follows the same general outline as replication in prokaryotes, with the most important difference being the presence of histone proteins complexed to eukaryotic DNA. 

EXAMPLE 8.7 One way in which difference 1 in Example 8.6 is addressed is that, in contrast to the single initiation site for DNA replication in prokaryotes, there are multiple initiation sites, between 3 x 10 and 3 x 10 base pairs apart, on eukaryotic chromosomes. Therefore, even though replication fork movement is slower in eukaryotes than in prokaryotes (about 50 nucleotides s ), the presence of multiple sites of initiation allows chromosome replication to occur on a time scale of 10 h whereas it would take -500 h if there were only a single initiation site. 

The enzymology of DNA replication in prokaryotes 227 TABLE 18.1 The subunit structure of DNA polymerase III holoenzyme of E. coli... 

In terms of evolutionary biology, the complex mitotic process of higher animals and plants has evolved through a progression of steps from simple prokaryotic fission sequences. In prokaryotic cells, the two copies of replicated chromosomes become attached to specialized regions of the cell membrane and are separated by the slow intrusion of the membrane between them. In many primitive eukaryotes, the nuclear membrane participates in a similar process and remains intact the spindle microtubules are extranuclear but may indent the nuclear membrane to form parallel channels. In yeasts and diatoms, the nuclear membrane also remains intact, an intranuclear polar spindle forms and attaches at each pole to the nuclear envelope, and a single kinetochore microtubule moves each chromosome to a pole. In the cells of higher animals and plants, the mitotic spindle starts to form outside of the nucleus, the nuclear envelope breaks down, and the spindle microtubules are captured by chromosomes (Kubai, 1975 Heath, 1980 Alberts et al., 1989). 

DNA plasmid-based treatment ( gene therapy ) is considered an alternative to the one based on classical chemical drugs or proteins recovered from recombinant cells. Treatment of acquired and inherent genetic diseases as well as the use of DNA for the purpose of vaccination are potential applications of plasmid DNA (pDNA). The plasmid carries information that allows protein expression in the targeted human cells as well as eukaryotic regulatory elements and specific prokaryotic sequences that control replication in the host cell, see Fig. 10. Formulation is required for ex- or in-vivo administration. Selected systems for gene expression can be viral or non-viral. 

Plasmid A relatively (cf. the chromosome) small (>20 Kb), usually circular, double-stranded DNA molecule found in prokaryotes capable of replicating independently of the chromosome. Plasmids carry genes which are usually not essential for the growth of the organism except under special conditions. Some plasmids carry genes for antibiotic resistance. See also Ti-plasmid. Some plasmids however can be very large, e.g. the plasmids in Rhizobium species. 

A. Replication-Associated Strand Asymmetries in Prokaryotic Genomes The Replicon model... 

DNA replication begins at the origin of replication (one in prokaryotes, multiple in eukary otes). The strands are separated locally, forming two replication forks. Replication of double-stranded DNA is bidirectional. 

Many important details have emerged concerning the mechanisms of DNA replication in both bacteria (Prokaryotes) and higher cells (Eukaryotes). These mechanisms are vital in understanding how a cell duplicates its genetic material (DNA), and how this duplication is related to cell division. For these reasons, cells have evolved elaborate mechanisms to ensure that the process of duplication (DNA replication) is error free. This level of control is so important that cells will actually cease cell division if errors become too frequent and wait until the DNA is repaired. 

TWn relatively recent developments have added to our knowledge significantly concerning how DNA replication occurs with fidelity or in what molecular biologists and biochemists call a processive polymerase activity. DNA polymerase is the enzyme which actually polymerizes (adds DNA precursors or building blocks) DNA. There are many such DNA polymerases in pro- and eukaryotic cells that have different functions but the main enzyme in prokaryotes is DNA polymerase 111 and in Eukaryotes. DNA polymerases alpha, delta, and epsilon. All four of these DNA polymerases are made of subunits. 

Viral Replication. In contrast to eukaryotic and prokaryotic cells, which multiply by binary fission, viruses multiply by synthesis of their separate components, followed hy assembly. Several stages aie involved in viral replication ... 

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