Gene Expresssion

MIC210
BASIC MOLECULAR BIOLOGY

Lecture 3
Gene Expression
By
SITI NORAZURA JAMAL (MISS AZURA)
03 006/06 483 2132

Outline
1. Gene expression in prokaryotic cells –
DNA to mRNA to protein.
2. Gene expression in eukaryotic cellsIntron splicing, 5’ capping, 3’-poly-A tail
3. DNA Replication
4. Reverse transcription

Every cell has the same DNA and therefore the same genes. But
different genes need to be “on” and “off” in different types of cells.
Therefore, gene expression must be regulated.

Gene expression must be regulated in
several different dimensions—
In time:

6 mos

14 wks

1 day

12 mos

10 wks

18 mos

At different stages of the life cycle, different genes need to be on and off.

1) Gene expression in
prokaryotic cells– DNA to mRNA
to protein.

1. Gene expression : DNA to mRNA to
protein
• Gene expression – process where the information in a gene is
read and used to synthesize a protein
• Genetic information is linearly transferred from DNA to protein.
• What proteins you can make depends on what genes you have
Gene expression in prokaryotes

Transcription
• a messenger RNA (mRNA) molecule is synthesize using
the antisense strand as a template
• the genetic information is now transferred to the mRNA
• RNA is like DNA except : ribose sugar, single stranded,
uracil

Molecular Components of Transcription
• RNA synthesis is catalyzed by RNA polymerase, which pries
the DNA strands apart and hooks together the RNA
nucleotides
• RNA synthesis follows the same base-pairing rules as DNA,
except uracil substitutes for thymine
• The DNA sequence where RNA polymerase attaches is called
the promoter; in bacteria, the sequence signaling the end of
transcription is called the terminator
• The stretch of DNA that is transcribed is called a transcription
unit

Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings

Translation
• the information in the mRNA is read in a set of 3 bases – a
codon
• each codon codes for an amino acid
• a chain of amino acids – a polypeptide – is built by reading the
codons
• all these happen in the ribosome in the cytoplasm

2) Gene expression in eukaryotic
cells- Intron splicing, 5’ capping,
3’-poly-A tail

The story is much more complicated in Eukaryotes

Important
differences
• A ‘cap’ is added to
the 5’ end of the
mRNA

• A polyA tail is
added to the 3’end
• Introns are
removed by a
process called
splicing

Introns and mRNA splicing
• Most eukaryotic genes
contain introns and exons
• Exons are DNA sequences
that carry genetic
information
• Introns do not carry genetic
information
• the introns are removed
from the mRNA by a
process called splicing
• whereby the introns are cut
out – and the exons are
rejoined
• this mature mRNA is then
translated to make proteins

3. DNA replication
Every new cell must have a complete set of genes
Before cell division occurs, the DNA is replicated so that each
new cell has its own set of DNA

Overview

Synthesis of
the leading
strand
during
DNA
replication

Origin of replication
Leading strand
Lagging strand

Primer
Lagging strand
Leading strand
Overall directions
of replication


3
5
RNA primer

5

“Sliding clamp”

3
5

Parental DNA

DNA poll III

3
5

5
3

5

In general :
• the original DNA molecules to serve as a template
• the new DNA strand is synthesized by the enzyme DNA
polymerase III
• Complementary base pairing ensures that the sequence of the
template is copied accurately

3‟

T

T

T

5‟
DNA polymerase III

New DNA strand

Synthesis of new DNA strand requires a primer
and can proceed only in a 5’  3’ direction (why?)

5’ PO4

In the cell, the primer is a short RNA molecule
In the test tube, a short piece of DNA will also work.

DNA replication step-by-step

1) Double helix structure opened up by a helicase enzyme
The single stranded regions are stabilized by SSBs (single
stranded binding proteins)


2) Another enzyme, primase, makes a short RNA
primer


3) Then DNA polymerase III begins to extend the new DNA
strand


Direction of
replicasome

4) All these enzymes work together in a complex known as a
replicasome.
The replicasome moves in one direction, following the replication
fork

The two strands of a DNA are not equal
(when it comes to replication)

Replication can only happen
in a 5‟ to 3‟ direction
„leading‟ and „lagging‟
strands


On the leading strand, everything’s OK

- DNA synthesis occurs continuously in a 5’  3’ direction


On the lagging strand, we have a problem
- DNA synthesis cannot happen in a 3’  5’ direction
- thus, multiple primers are made
- new DNA is synthesized as small Okazaki fragments (5’  3’)
- the primers are then replaced with DNA by DNA polymerase I
- the DNA fragments are then joined by DNA ligase

Fig. 16-17

A summary of bacterial DNA replication
Overview
Lagging strand
Leading strand

Leading strand
Lagging strand
Overall directions
of replication

Single-strand
binding protein
Helicase

5
3
Parental DNA

Leading strand

3

DNA pol III
Primer
5

Primase
3
DNA pol III

Lagging strand

5
4

DNA pol I

3 5
3

2

DNA ligase
1

3
5

3

Synthesis of the
lagging strand

5
5

Template
strand

3

3

RNA primer

3

1

5

5

3

1

5
3

5

2

3

3
5

Okazaki
fragment

3
5

1

5
3
5
2

1

5
3
1
2

Overall direction of replication

3
5

Proof reading minimized replication error

DNA polymerase III has a 3‟  5‟ exonuclease activity that can
cut and repair mistakes
Remember : DNA replication has to be very accurate (or else?)

DNA replication is semi conservative

Replication : From one DNA molecules to two
Identical sequences

4. Reverse transcription – from RNA to DNA

The transfer of genetic information from RNA to DNA
• By the enzyme reverse transcriptase found in retrovirus
• This allows us to make cDNA (complementary DNA) from mRNA
• and obtain a gene sequence without the introns

Reverse transcription

cDNA

Gene Expresssion

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