DNA Replication: Mechanisms, Enzymes, and E. Coli Process
DNA Replication Fundamentals
The organism has the ability to create exact copies of itself, based on a template. This was discovered when it was clear that DNA somehow acts to serve as a template in replication and transcription of genetic information. Each of the two strands are complementary to each other, following these rules:
- DNA replication is semiconservative: Each DNA strand is the template for the synthesis of a new chain. The result is two new DNA molecules, each with a new chain and an old one.
- Replication begins at a point of origin and progresses bidirectionally.
- DNA synthesis progresses in the 5′-3′ direction. One of the new chains is synthesized in the form of short pieces called Okazaki fragments, meaning one strand is synthesized continuously and one discontinuously.
Enzymes that degrade DNA are called nucleases. Exonucleases degrade nucleic acids from the molecule’s extremity, while endonucleases start at specific internal sites.
- All require a DNA template.
- A primer with a free 3′ hydroxyl group is necessary for nucleotides to be added.
- DNA replication requires a special degree of fidelity. Internal mechanisms, such as exonuclease activity, perform a second check after each nucleotide is added.
E. Coli Replication
Over 90% of E. coli DNA polymerase is developed by DNA polymerase I. While it may not be a suitable enzyme for chromosome replication, the search for polymerases led to the discovery of DNA polymerase II (involved in DNA repair) and DNA polymerase III (the primary enzyme in E. coli replication).
DNA polymerase I performs multiple functions, including cleaning during replication. DNA polymerase III is more complex, with polymerization and proofreading activities residing in the alpha and epsilon subunits.
E. coli polymerase replication needs more than 20 different enzymes and proteins. Separation of the two parental strands requires helicases, enzymes that unfold and move along DNA, utilizing ATP chemical energy to separate the chains, creating surface tension in the structure. Primers are synthesized by enzymes called primase and attach to the DNA before synthesis starts. Eventually, the DNA primer is replaced.
DNAA is the key component that recognizes and successively denatures DNA from the region of the 3 repetitions of the 13 base pair, which are rich in AT. DNA B binds to the region where the DNA. Two hexamers of DNA B act as helicases, unwinding DNA and creating two forks, while a gyrase relieves surface tension.
Elongation
Elongation involves two operations: continuous chain synthesis and lagging chain synthesis. From this point, the synthesis of both strands differs.
Termination
The two replication forks meet on the E. coli chromosome in a terminal region containing multiple copies of a 20 base pair sequence, the binding site for TUS protein, which can stop the replication fork.