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10.2.5.1. Replication in prokaryotes

DNA replication in prokaryotes


Initiation

The replication begins a sequence of nucleotides in the DNA called origin of replication , oriC or initiation point, which acts as an initiation signal. This sequence is different depending on the species, but it has abundant thymine (T) and adenine (A). The T and A are linked by two hydrogen bonds, instead of three, like the C and G, so these bonds will be weaker and easier to break.

The following proteins are involved in the initiation of replication:

  • Helicasas. They are enzymes that recognize the nucleotide sequence origin of replication and break hydrogen bonds between complementary nitrogenous bases. They are responsible for opening the double helix so that the chains can serve as a mold for the new chains.


  • Topoisomerases. The unwinding of the double helix creates stresses between the two strands, and the topoisomerases are responsible for cutting the strands to release the supercoiling stresses. Cutting a (the topoisomerases I ) or both chains (the topoisomerase II or gyrase) of DNA, and when there are no such tensions, ligases the spliced again.


  • SSB proteins (Single Strand Binding-DNA). They are the stabilizing proteins that bind to each strand of DNA separated by the helicase so that they do not rejoin. Thus, they allow the correct passage of DNA polymerase , preventing complementary strands from joining before the nucleotides of the new strand that is being formed are added.
    • helicase works in each direction, so this process is bi-directionalThe two replication forks that have been created form the replication bubbles or eyes .

      As DNA polymerase needs to have a primer to which nucleotides can be added, an RNA polymerase must first intervene that synthesizes a small fragment of about ten nucleotides of RNA that serves as a primer. To this RNA polymerase it is called primase, and a fragment of RNA that serves as a primer, first.

    Elongation (formation of new strands)

    The DNA polymerases will be responsible for initiating the synthesis of complementary daughter strands.

    In each replication fork, the chain forward and delayed grow differently:

    Synthesis of the leading or leading chain

    This chain is complementary to the chain 3'→ 5'. After the RNA polymerase has synthesized the first, primer RNA, DNA polymerase III, it begins to synthesize the new strand in the 5'→ 3' direction. The energy needed for this process is provided by nucleotides, which lose one of their phosphate groups.

    This new chain is of continuous growth, since the helicase does not stop.

    Synthesis of the delayed chain

    In the other complementary strand, the DNA polymerase should read the strand in the 5'→ 3' sense, adding nucleotides to the new strand in the 3'→ 5' sense, which is not possible.

    This new chain is of discontinuous growth, from separated DNA fragments. It is called a delayed chain or strand because its synthesis is slower than that of the leading chain.

    Synthesis of this strand begins when RNA primase synthesizes about 40 nucleotides of RNA, the primer RNA, at a point that is about 1000 nucleotides away from the initiation signal.

    The DNA polymerase III binds in a 5'→ 3', about 1000 or 2000 nucleotides of DNA, forming an Okazaki fragment. This process is repeated as the two chains that serve as a mold separate.

    Hebra retardada: síntesis de cebadores, unión de fragmentos de Okazaki y eliminación de los cebadores.

    By César Benito Jiménez [CC BY-SA 2.5 es or CC BY-SA 2.5 es], via Wikimedia Commons

    Afterwards, DNA polymerase I, due to its exonuclease function, eliminates the primer RNAs, and later, due to its polymerase function, fills the gaps occupied by the ribonucleotides with DNA nucleotides.

    Finally, DNA-ligase joins the different Okazaki fragments with a phosphodiester bond.

    The DNA strands that serve as templates are also called parental DNA.

    By César Benito Jiménez [CC BY-SA 2.5 es or CC BY-SA 2.5 es], via Wikimedia Commons

    Termination

    The elongation is continued until the DNA is completely replicated. As the growth of the chains is bidirectional, one of the new strands has been synthesized continuously and the other discontinuously, using the Okazaki fragments. The two replication forks will join in a place diametrically opposite to the origin of replication of the bacterial chromosome .

    The DNA polymerase I remove the last primer, and the fragments are joined by DNA ligase. Thus, two strands of DNA are obtained, without RNA, and identical to the parental DNA molecules.

    Fundamental insights on DNA replication in prokaryotes

    DNA replication in prokaryotes

    • Initiation.
      • It begins in the DNA sequence called the origin of replication.
      • Enzymes involved:
        • Helicasas.
        • Topoisomerases.
        • Stabilizing proteins SSB.
      • Bidirectional replication. Replication forks are formed.
      • The RNA polymerase (primase) synthesized a 10 nucleotide RNA which serve as primer (PRIMER) . 
    • Elongation (formation of new strands).
      • Synthesis of the leading or leading chain.
        • It is complementary to the chain 3'→ 5'. 
        • The DNA polymerase III, begins to synthesize the new chain in the 5'→ 3', after the RNA polymerase is synthesized first, RNA primer.
        • Continuous growth.
      • Synthesis of the delayed chain.
        • Discontinuous growth, slower than the leading chain.
        • The primase synthesizes RNA primer, and the DNA polimesa III nucleotide added in the 5'→ 3' forming an Okazaki fragment.
        • The DNA polymerase I removes the RNA primer and complete the gaps with nucleotides of DNA.
        • The DNA ligase joins a phosphodiester bond different Okazaki fragments.
    • Termination.
      • The RNA polymerase I removes the last primer and the fragments are joined by DNA ligase.