DNA replication involves the synthesis of new DNA strands through a semi-conservative process whereby each new molecule contains one old strand and one new strand synthesized using the old strand as a template. Key enzymes involved include helicase, which unwinds the DNA double helix, primase, which initiates DNA synthesis, and DNA polymerase, which catalyzes phosphodiester bond formation to elongate the new strands. Fidelity is maintained through proofreading mechanisms that remove incorrectly incorporated nucleotides and DNA repair pathways that correct errors made during replication.
1. Fidelity of DNA replication
INDRANI KAR
M.Sc Microbiology
St. George College of Management & Science
Bengaluru North University
2. Introduction
• The metabolism of DNA is the chemistry of joining one nucleotides to the next in the
process called DNA replication, is elegant and simple.
• The metabolism of DNA or the DNA replication needs high accuracy that is arrived
by enzymes complexity.
• The enzymes that synthesize DNA molecules are very well known for their ability in
each work in replication, and it may attribute the copies of millions of bases.
• It has done by extraordinary fidelity and speed with the formation of
phosphodiester bonds in proper manner.
3.
4. History of replication
• Watson and Crick proposed the hypothesis of the DNA replication by the model of semi-
conservative model of replication.
• Each DNA strands serves as the template strands for synthesis of a new strand, producing
two new DNA molecules, each with new strand and one old strand. This is semi-
conservative replication.
• After the promising hypothesis of semi-conservative replication, in 1957 Meselson and
stahl grew E. coli cells for generation in the medium in which the sole nitrogen source
NH4CL containing heavy and light isotopes of nitrogen.
• They proved that it is DNA replication is a semi conservative mode of replication , the
DNA possess a new daughter strand and a old mother strand.
5.
6. Enzymes for replication
• Single stranded binding protein or enzyme.
• Helicase enzyme.
• Primase enzyme.
• DNA polymerase III
• DNA polymerase I
• DNA ligase
• DNA gyrase (DNA topoisomerase II)
• Dam methylase enzyme
7.
8. Proteins for replication
• DnaA protein
• DnaB protein
• HU (Histone like bacterial protein)
• FIS ( factor for inversions stimulation )
• IHF (integration host factor)
• DnaG protein
9. Initiation of DNA replication
• The E.coli replication origin oriC , consists of 245 bp and contains DNA sequence
elements that are highly conserved among replication origins.
• DNA first unwinds at the position of A=T base pairs called the DNA unwinding
element.
• DNA unwinds by the DNA unwinding protein or DnaA , DnaB , DnaC and these
three proteins are called helicase enzyme.
• When DNA unwinds the single stranded binding protein stabilized the unwind
DNA single strand.
11. Elongating of replication
• The elongation phase of replication includes two distinct but related operations by
adding same nucleotides.
• At first the primase enzymes add small small nucleotides at the two strands, it only
synthesize the small fragments like 9 to 10.
• When the small fragments are replicated by primase then the polymerase III replicates
long nucleotides by forming the okazaki fragments
• Okazaki fragments leading to the synthesis of two strands leading strand and lagging
strand as in the oposite strand.
• The processivity of DNA replication is done by topoisomerase enzymes.
12.
13.
14. Termination of replication
• Eventually the two replication fork of the circular chromosome meet at the
termination region.
• The termination region containing 20 bp sequence called Ter sequence.
• The tar region arranged on the chromosome to create a trap that a replication fork,
the ter sequence function as binding sites for the protein tus.
• The tus-ter complex can arrest the replication fork from one direction.
• The replication fork encounters a function tus-ter complex, it halts, and other halts
when it meets the first fork.
• Separation of DNA strands are done by topoisomerase, the two daughter strand or
cells segregated by cell division.
16. Fidelity of DNA replication
• Fidelity of DNA replication is depend on the enzyme to catalyze many
reactions before relasing its substrate is called processivity.
• The key of processivity of DNA polymerase that act at the replication
fork in their association with proteins called sliding DNA clamps.
• The speed of DNA synthesis is largely due to the progressive nature of
DNA polymerase III due to it binds with sliding DNA clamps and stabilizes
itself in elongation process.
• Thus the replication fidelity increases.
17.
18. Continue......
• The another reason for fidelity is the proof reading process of replication.
• In proof reading process, the incorrect base pairs remove by the nucleus that originally
identified in the same polypeptide as the DNA polymerase.
• In the polymerase 1 two nuclease , one is exonuclease and one is endonuclease.
• The two nuclease form a functional large fragments called klenow fragment, retains the
polymerization and proof reading activity.
• Thus the fidelity is increased of the DNA replication.
20. CONCLUSION
• DNA replication itself can occasionally damage, called mutation that introduces
error, hence the maintainance of the DNA replication is needed for the next
generation purpose.
• DNA molecules are themselves irreplaceable, it can cause cellular damage too.
• Fidelity of DNA replication depends on the proof reading, repair system as well
as in some enzyme 's function.
• Such mechanism of DNA replication fidelity is also depends on the onway repair
system like nick translation done by the phosphodiester bonds and with the help
of ATPs.
