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In prokaryotes, DNA polymerase I synthesizes the Okazaki fragment until it reaches the previous RNA primer. Then the enzyme simultaneously acts as a 5′→3′ exonuclease, removing primer ribonucleotides in front and adding deoxyribonucleotides behind.
After replication of the desired region, the RNA primer is removed by DNA polymerase I via the process of nick translation. The removal of the RNA primer allows DNA ligase to ligate the DNA-DNA nick between the new fragment and the previous strand. DNA polymerase I & III, along with many other enzymes are all required for the high fidelity ...
DNA polymerase I (or Pol I) is an enzyme that participates in the process of prokaryotic DNA replication. Discovered by Arthur Kornberg in 1956, [1] it was the first known DNA polymerase (and the first known of any kind of polymerase). It was initially characterized in E. coli and is ubiquitous in prokaryotes.
There are two main types of primase: DnaG found in most bacteria, and the AEP (Archaeo-Eukaryote Primase) superfamily found in archaean and eukaryotic primases. While bacterial primases (DnaG-type) are composed of a single protein unit (a monomer) and synthesize RNA primers, AEP primases are usually composed of two different primase units (a heterodimer) and synthesize two-part primers with ...
The E. Coli DnaG primase is a 581 residue monomeric protein with three functional domains, according to proteolysis studies. There is an N-terminal Zinc-binding domain (residues 1–110) where a zinc ion is tetrahedrally coordinated between one histidine and three cysteine residues, which plays a role in recognizing sequence specific DNA binding sites.
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m4C methyltransferases (N-4 cytosine-specific DNA methylases) are enzymes that specifically methylate the amino group at the C-4 position of cytosines in DNA. [5] Such enzymes are found as components of type II restriction-modification systems in prokaryotes. Such enzymes recognise a specific sequence in DNA and methylate a cytosine in that ...
Use of the thermostable Taq enables running the PCR at high temperature (~60 °C and above), which facilitates high specificity of the primers and reduces the production of nonspecific products, such as primer dimer. Also, use of a thermostable polymerase eliminates the need to add new enzyme to each round of thermocycling.