DNA Replication in Prokaryotes

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Introduction

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DNA replication has turned out to be a much more complex process than we originally thought. Some of this complexity arises because of two reasons which you should keep in mind:

1. The DNA molecule is antiparallel, meaning that the strand or halves of the molecule are oriented in opposite directions. In this diagram the left strand (reading here from top to bottom) is the 5'---> 3' strand and the right strand is the 3'---> 5' strand.

2. The enzymes which lay down the complementary strand to each parent strand only travels in a 3' ---> 5' direction relative to the parent strand. 

So DNA synthesis in the left hand strand of my diagram would be from bottom to top of the page, whereas DNA synthesis using the right hand strand would be from top to bottom since for the right hand strand this corresponds to the 3' --->5' direction.

Note that the new strand itself is growing from its 5' end to its 3' end. 

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Steps in DNA Replication in Prokaryotes: E. coli

Initiation

1. Original DNA molecule. The parent strands and are shown in red.

2. Initiator proteins attach to a specific initiation site, recognized as a particular sequence of DNA nucleotides.
3. DNA Helicase attaches at initiation site. Other proteins called SSB(single strand binding proteins), prevent reannealing of the DNA.

DNA Helicase is an enzyme that unravels the DNA double helix and breaks the hydrogen bonds.

4. Replication fork formation.

Replication fork. The replication fork refers to the region on the DNA that is the leading edge of DNA replication as the DNA unzips in a particular region. At this stage the replication fork has a complex of proteins including the enzyme helicase and another enzyme, DNA primase.

DNA primase is an enzyme that generates an RNA sequence that serves as a starting point or primer for synthesis of the new DNA chain.

 

5. Start of DNA synthesis on leading strand of DNA using DNA polymerases. 

DNA polymerases are enzymes that synthesize a complementary DNA strand using a the original strand as a template.  In DNA synthesis the new strand grows 5' to 3'.

Elongation

1. Production of new DNA half from parental leading strand as a template. Remember that DNA synthesis is in 3' --> 5' direction of the parental strand. Leading strand synthesis is continuous from the primer to the replication fork. 

Meanwhile another RNA primase(Light brown) has attached an RNA primer(Purple) to the upper parental strand. This other new strand is the lagging strand. It's synthesis is discontinuous.

2. Production of first Okazaki fragment in the lagging strand

An Okazaki fragment(a) is a stretch of non parental DNA produced along the lagging strand of parental DNA by the DNA polymerase beginning at primer(b). Since the initiator proteins do not move at this stage, the DNA polymerase has to stop synthesis when it is encountered. Primer(c) is shown being synthesized by the DNA primase.

3. Production of of more Okazaki fragments and joining of fragments using ligase.

Synthesis of DNA along the leading strand still continues and multiple Okazaki fragments are produced by DNA polymerase III. Once several fragments are produced, the primer is removed by another DNA polymerase called DNA polymerase I 1 (b). Then another enzyme, a ligase joins the DNA fragments together (a). 

This process continues until all the DNA is replicated.

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pgd. Created 08/01/2002 revised 10/20/2005
 
 

Bi-directional synthesis

In many prokaryotes two replication forks develop from at the initiation site, each initiation fork traveling in opposite directions as shown here. Thus, what strand is lagging or leading depends on which replication fork you are dealing with!