Lecture 6: DNA replication

By: Dr samar elkhateeb

lecturer of biochemistry and genetic
[email protected]
 DNA replication (duplication)

• Replication of DNA occurs during the process of normal cell division cycle as it is

responsible for cell reproduction prior to cell division.

• The entire process of DNA replication is complex and involves multiple enzymatic

activities.

• The rate of replication in humans , about 40 to 50 nucleotides per second.

• The process of DNA replication begins at specific sites termed origins of replication,
DNA replication (duplication)

• DNA replication is unable to reach the very end of the chromosomes, but

ends at the telomere region of repetitive DNA close to the end.

• This shortens the telomere of the daughter DNA strand. Shortening of the

telomeres is a normal process in somatic cells.

• As a result, cells can only divide a certain number of times before the

DNA loss prevents further division. This is known as the Hayflick limit.
 Cont.

• DNA polymerase elongate a new DNA strand by adding nucleotides to the

3` end of the growing chain ,so the new strand grows in 5`to 3` direction

• In the last step DNA polymerase removes the RNA primer and replace it
with deoxynucleotides, the sections are joined by the action of an enzyme
called DNA ligase.
Separation of the 2 strands 1 Formation of the replication fork 2
The DNA double helix • Area where the 2 strands
unwinds and hydrogen unwind they form a “V” where
bonds joining the base pairs active synthesis occurs.
break this is done by an • Single-stranded DNA binding
enzyme known as proteins (SSB) assure that the
DNA helicase parental DNA strands will not
re anneal together,

DNA polymerase
Topoisomerase enzyme
• cannot begin synthesis with a
• Prevents entangling of DNA single nucleotide,
stands during the process of • requires a free OH as a
unwinding substrate for ELONGATION.

Primase
• Synthesizes a short stretch of
RNA complementary to DNA
(primer)
Steps of replication

Separation of the 2 strands:

• In order for the 2 strands of the parental double helical DNA to be
replicated, they must separate.

• The DNA double helix unwinds and hydrogen bonds joining the base pairs
break this is done by an enzyme known as DNA helicase.
Cont.

Formation of the replication fork:

• As the 2 strands unwind they form a “V” where active synthesis occurs ,
this area is called the replication fork.

• Single-stranded DNA binding proteins (SSB) assure that the parental DNA
strands will not re anneal together, thus giving the chance for replication via
complementary base paring
• Moreover, topoisomerase enzyme acts to prevent entangling of DNA
stands during the process of unwinding.
• DNA polymerase enzyme cannot begin synthesis with a single
nucleotide , instead DNA polymerase enzyme requires a free OH as a
substrate for ELONGATION.
• Primase synthesizes a short stretch of RNA complementary to DNA
(PRIMER).
The leading strand
• it is the strand that is copied in the direction towards the replication
fork and it is synthesized continuously.

• synthesis of DNA proceeds in the 5' to 3' direction through the

attachment of the 5'-phosphate of an incoming (deoxytriphosphate)
dNTP to the existing 3'-OH in the elongating DNA strands with the
release of pyrophosphate.
lagging strand
• It is the strand that is being copied away from the replication fork and is
synthesized discontinuously.

• The short segments of this discontinuous DNA are called Okazaki

fragments, they are then joined together to form continuous strand
Gene expression
Genome

• It is the complete set of information in an organism’s DNA.

• Total length of DNA about 2 meters long in a human cell, encoding

about 30000 proteins.

Gene

• It is the total sequence of nitrogenous bases in DNA that specifies

the amino acid sequence of a protein molecule. 15

Gene structure

The gene consists of:

• Promoter (where RNA polymerase binds) which is important for initiation of

transcription.

• Exons (the coding sequence).

• Introns (the non coding sequence).

• Start codon (the first codon of the gene ).

• Stop codon ( the end of the gene).

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• Terminator ( where the transcription ends).
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Gene expression (Protein synthesis)

 Protein synthesis pass through 2 main process.

1. Transcription

2. Translation

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Transcription

 It the process by which mRNA is formed from a DNA template .

 RNA polymerase binds to the promoter site on DNA (nucleotide sequence
which includes start point).
 It also, determines which DNA strand will be used as a template (antisense
).
 The antisense strand will act as a guide for selection of its complementary
nucleotides.
i. e T will select A, A will select U

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Cont.

• Once transcription begins the RNA polymerase moves along the DNA and
opens a short segment of DNA, assembling the complementary mRNA strand
from 5` to 3`.

• The general steps required to synthesize the primary RNA transcript are
initiation, elongation, and termination.

• Heterogeneous nuclear RNA (HnRNA): pre-mRNA which exists only before it

is fully processed into mRNA. 20
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• The mRNA strand consists of coding regions called exons separated by regions
which don't contribute in protein synthesis called introns.

• After transcription, the introns in the precursor mRNA are excised, the exons
are spliced together to form mature m RNA .

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Cont.

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Post transcriptional processing

• RNA transcripts are subjected to post –transcription modifications as they are

capped at the 5` end by addition of modified methyl guanine this capping is
important to prevent RNA molecule from degradation.

• At the 3` poly A tail is added , which may be involved in stabilizing the mRNA
not to be degraded.
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Translation

• mRNA leaves the nucleus to the cytoplasm where It associates with ribosomes .

• Amino acids in the cytoplasm are activated by ATP and get linked to tRNA, each tRNA molecule

carries 3 bases complementary to a specific codon on mRNA (anticodon).

• The amino acid tRNA complex then moves to ribosomes to be arranged in order by base

pairing between a codon on mRNA and the corresponding anticodon on tRNA.

• Ribosomes then move along the mRNA while amino acids are linked together forming the
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 Gene Regulation
• Different cells within an organism share the same set of chromosomes, in each
cell some genes are active while others are not

• Each cell produces different proteins according to its needs so that it does not
waste energy by producing proteins that will not be used

• Certain genes are found in order to regulate the protein synthesis, these genes
are called regulatory genes,

• an operon is a functioning unit of genomic DNA containing a cluster of

genes under the control of a single regulatory signal or promoter 27
Biomedical Importance

• It was impossible to understand protein synthesis or to explain mutations

before the genetic code was elucidated.

• The code provides an explanation for the way in which protein defect may
cause genetic disease and for the diagnosis and perhaps treatments of these
disorders.

• In addition the pathophysiology of many viral infections is related to the ability

of this agents to disrupt host cell protein synthesis. 28