Lecture 7

Molecular Biology ٣٢٥ Sabah Linjawi ١

 DNA replication
„ DNA replication begins with the "unzipping" of the parent molecule as the
hydrogen bonds between the base pairs are broken.
„ Once exposed, the sequence of bases on each of the separated strands serves
as a template to guide the insertion of a complementary set of bases on the
strand being synthesized.
„ The new strands are assembled from deoxynucleoside triphosphates.
„ Each incoming nucleotide is covalently linked to the "free" 3' carbon atom on the
pentose as
„ the second and third phosphates are removed together as a molecule of
pyrophosphate (PPi).
„ The nucleotides are assembled in the order that complements the order of
bases on the strand serving as the template.
„ Thus each C on the template guides the insertion of a G on the new strand,
each G a C, and so on.
„ When the process is complete, two DNA molecules have been formed identical
to each other and to the parent molecule.

Molecular Biology ٣٢٥ Sabah Linjawi ٢

 DNA replication

„ The double helix is

unwound and each
strand acts as a
template.
„ Bases are matched to
synthesize the new
partner strands

Molecular Biology ٣٢٥ Sabah Linjawi ٣

 Semi-conservative mechanism

„ This process is "semiconservative" in that each

strand of the original double-stranded DNA molecule
serves as template for the reproduction of the
complementary strand.
„ following DNA replication, two identical DNA
molecules have been produced from a single
double-stranded DNA molecule.
„ Cellular proofreading and error-checking
mechanisms is used in genetics to refer to the
error-correcting processes involved in DNA
replication

Molecular Biology ٣٢٥ Sabah Linjawi ٤

 Types of DNA replication

Molecular Biology ٣٢٥ Sabah Linjawi ٥

 Replicons, origins and termini

„ Small chromosomes, such as those of bacteria and

viruses, replicate as single units called replicons.
„ Replication begins from a unique site, the origin,
and proceeds, usually bidirctionally, to the terminus.
„ Large eukaryotic chromosomes contain multiple
replicons, each with its own origin, which fuse as
they replicate
„ Origins tend to be AT-rich to make opening easier

Molecular Biology ٣٢٥ Sabah Linjawi ٦

 Replicons, origins and termini

Molecular Biology ٣٢٥ Sabah Linjawi ٧

 Semi-discontinuous replication
„ At each replication fork, the leading strand is synthesized as one
continuous piece
„ While the lagging strand is made discontinuously as short
fragments in the reverse direction
„ The leading strand is that strand of the DNA double helix that is
orientated in a 5' to 3' manner
„ An Okazaki fragment is a relatively short fragment of DNA (with
an RNA primer at the 5' terminus) created on the lagging strand
during DNA replication.
„ The lengths of Okazaki fragments are between 1,000 to 2,000
nucleotides long in E. coli and are generally between 100 to 200
nucleotides long in eukaryotes.

Molecular Biology ٣٢٥ Sabah Linjawi ٨

 Semi-discontinuous replication

Molecular Biology ٣٢٥ Sabah Linjawi ٩

 RNA priming
„ A primer is a strand of nucleic acid that serves as a starting point
for DNA replication.
„ They are required because the enzymes that catalyze replication,
DNA polymerases, can only add new nucleotides to an existing
strand of DNA.
„ The polymerase starts replication at the 3'-end of the primer,
and copies the opposite strand.
„ In most cases of natural DNA replication, the primer for DNA
synthesis and replication is a short strand of RNA.
„ This RNA is produced by primase, and is later removed and
replaced with DNA by a repair polymerase.

Molecular Biology ٣٢٥ Sabah Linjawi ١٠

 Origins of Replication

„ Eukaryotic chromosomes have multiple origins of

replication
„ In yeast, these are known as Autonomously Replicating
Sequences (ARS).
„ Replication in eukaryotic cells must be coordinated
with the stages of the cell cycle.
„ DNA synthesis only occurs during S phase.
„ Mitosis occurs during the M phase.
„ These two phases are separated by two gaps: G1 and
G2.

Molecular Biology ٣٢٥ Sabah Linjawi ١١

 Replication cycle

Molecular Biology ٣٢٥ Sabah Linjawi ١٢

 The Enzymes
„ The eukaryotic cells have more than a dozen DNA polymerases.
„ Two of these enzymes (α and δ) are important for the replication of
eukaryotic chromosomes.
„ The rate of synthesis of DNA in eukaryotic cells is about one tenth
the rate of bacterial DNA synthesis
„ DNA polymerase alpha (α) is composed of 4 subunits, one of which
(167 kDa) carries the polymerase activity.
„ It is responsible for synthesis of the primer on the lagging strand.
„ The primer consists of both RNA and a short stretch of DNA.
„ DNA polymerase delta (δ) contains at least 4 and maybe as many
as a dozen subunits.
„ It has a proofreading activity.

Molecular Biology ٣٢٥ Sabah Linjawi ١٣

 Enzymes function

Molecular Biology ٣٢٥ Sabah Linjawi ١٤

 Replication fork
„ The replication fork is a structure that forms within the nucleus
during DNA replication
„ It is created by helicases ،which break the hydrogen bonds
holding the two DNA strands together.
„ The resulting structure has two branching, each one made up of
a single strand of DNA, that are called the leading and lagging
strands .
„ DNA polymerase creates new partners for the two strands by
adding nucleotides
„ Because DNA polymerase can only synthesize a new DNA
strand in a 5' to 3' manner, the process of replication goes
differently for the two strands comprising the DNA double helix.

Molecular Biology ٣٢٥ Sabah Linjawi ١٥

 The DNA replication fork. RNA primer
labeled at top

Molecular Biology ٣٢٥ Sabah Linjawi ١٦

 Telomere replication

„ Telomeric DNA consists of multiple copies of a simple

repeat sequence
„ The 3`- ends overhange the 5`- ends
„ Telomeric DNA is copied by telomerase, which carries
the template for repeat as a small RNA molecule
„ Telomerase is repressed in somatic cells but reactivated
in many cancer cells

Molecular Biology ٣٢٥ Sabah Linjawi ١٧

 A telomere or t-loop

Molecular Biology ٣٢٥ Sabah Linjawi ١٨

 References

„ users.rcn.com/jkimball.ma.ultranet/.../DNAReplication.ht
m
„ www.mun.ca/biochem/courses/3107/.../Euk_replication.
html
„ Molecular Biology. P.C. Turner, A.G. Mclennan, A.D.
Bates & M.R.H. White.School of Biological Sciences,
University of Liverpool, Liverpool, UK. Second edition.
BIOS Scientific Publishers, 2000.

Molecular Biology ٣٢٥ Sabah Linjawi ١٩