Meselson and Stahl's experiment provided evidence for the semi-conservative model of DNA replication proposed by Watson and Crick. DNA replication involves enzymes that unwind, separate, and stabilize the parental strands. DNA polymerase synthesizes new leading and lagging daughter strands while DNA ligase seals fragments. Replication follows the central dogma where DNA is transcribed to RNA and translated to protein.

1. Modes of replication; Meselson and Stahl’s
experiment Semi-conservative
replication; Okazaki fragments, enzymes
and proteins in DNA replication;
prokaryotic & eukaryotic DNA
polymerases; types and their functions
Presented by
Jyoti Devendra Adala
Msc part II

2. Transcription is the first stage in the expression of genetic information, code
contained in nucleotide sequence of mRNA molecules is translated thus
completing gene expression. This flow of information from DNA to RNA to
protein is termed as central dogma of molecular biology.
Replication is the process in which a parent DNA molecule forms two daughter
DNA molecules which are similar to itself. To know how DNA is replicated to
another DNA. Several models have been proposed. The three majors models are:-

4. Watson and crick proposed the hypothesis of semi conservative replication
and the hypothesis proven by Meselson and Stahl using DGC.

5. Enzymes and proteins involved in replication
DNA helicase breaks the hydrogen bonds
between the DNA strands.
Topoisomerase alleviates positive
supercoiling.
Single-strand binding proteins keep the
parental strands apart.
Primase synthesizes an RNA primer.
DNA polymerase III synthesizes a daughter
strand of DNA.
DNA polymerase I excises the RNA
primers and fills in with DNA (not shown).
DNA ligase covalently links the Okazaki
fragments together.

6. Primosome
It is a complex containing Pri A, Pri B, Pri C, Dna
T, Dna B, Dna C and Primase (Dna G) proteins.
Primososme complex without primase referred as
preprimosome complex.
Primosome complex plays a vital role in lagging
strand synthesis by synthesizing the primer at
frequent intervals.

7. DNA Helicases
There are four kinds of helicases:
DNA-A protein - It binds to 4 of 9 bp
sequence and moves in 3 of 13 bp sequence.
It is the first protein which binds to DNA to
initiate DNA replication.
DNA-B proteins - It is responsible for the
extension of open complex during
replication.
Rep Proteins - It is a helicase consisting of
one subunit. It binds to the 5'3' template.
DNA Helicase - II - It is helicase consisting
of only one subunit. It binds with 3'5'
template stand.

8. TOPOISOMERASES
Topoisomerase type II

9. Single-stranded DNA-binding
protein, or SSB
It was called a DNA-unwinding
protein. SSB binds to single-
stranded regions of DNA to
prevent premature annealing, to
protect the single-stranded DNA
from being digested by nucleases.
DNA primase:
DNA primase is an enzyme involved in the replication of DNA. DNA primase
is, in fact, a type of RNA polymerase which creates an RNA primer (A primer
is a strand of nucleic acid that serves as a starting point for DNA synthesis).

10. DNA POLYMERASE I
DNA Polymerase catalyze the formation of
polynucleotide chains through the addition of
successive nucleotides.
DNA polymerase I catalyzes formation of
phosphodiester bond between 3’-OH of the
deoxyribose and the 5’-phosphate of the dNTP.
DNA Polymerase has three activities
1) DNA polymerase activity
2) A 3’→5’ exonuclease activity that
hydrolyzes off mispaired nucleotide.
3) A 5’→3’ exonuclease activity that remove
the RNA primers.
DNA Polymerase II is more processive than
Pol I and has the same 3’ →5’ activity as
polymerase but lacks 5’→3’ exonuclease
activity

11. DNA polymerase III
DNA polymerase III begins synthesizing DNA in the 5’ 3’direction, beginning at the 3’
end of each RNA primer. The newly synthesized strand is complementary and antiparallel
to the parental strand used as a template.

12. DNA POLYMERASE-EUKARYOTIC
i) DNA polymerase α : It is composed of four subunits of which one has the
primase activity. DNA pol α is supposed to carryout lagging strand synthesis.
ii) DNA polymerase β: It is involved in the repair of DNA.
iii) DNA polymerase ϒ: This polymerase is involved in the replication of
mitochondria DNA.
iv) DNA polymerase δ: This polymerase has polymerase function and 3'-->5’
exonuclease activity. It is involved in leading strand synthesis.
v) DNA polymerase Ɛ: It is supposed to replace DNA pol delta in some
situations such as DNA repair.
vi) DNA polymerase σ: This polymerase seems to be expressed only in the
bone marrow cells. It is the only eukaryotic polymerase that has a
deoxyribonuclease activity.

13. DNA LIGASE:- DNA ligase seals the "nicks" between Okazaki fragments,
converting them to a continuous strand of DNA.
OKAZAKI FRAGMENTS:- Fragments synthesized during lagging strand formation
of replication was identified and proved by Rejis Okazaki.

14. REFERENCES
• MOLECULAR BIOLOGY BY DAVID CLARK
• BIOCHEMISTRY BY L STRYER, J M BERG AND J L TYMOCZKO
• MOLECULAR AND CELL BIOLOGY 5TH EDITION BY LODISH
• BIOCHEMISTRY BY LIPPINCOTT