Molecular

Biology
&
Genetic
Engineering
2024/2025
By Abdallah Noui
DNA Transcription
and translation
Protein synthesis
=
Gene expression
The Central Dogma
of Molecular Biology

Cell

Transcription DNA

mRNA
Translation Ribosome

Polypeptide
(protein)
©1998 Timothy G. Standish
 Proteins
• Proteins are made up of a chain of amino acids.
• Proteins are enzymes, which catalyze and
regulate chemical reactions.
 DNA Structure
The building blocks of DNA are
called Nucleotides.

One nucleotide is made of 3

important things:
1. 5-Carbon Sugar Deoxyribose
2. Phosphate
3. Nitrogen base
there are 4 nitrogen bases in
DNA: Adenine, Guanine,
Cytosine, and Thymine that
pair together)

AT C G
 RNA Structure
The building blocks of RNA are
Nucleotides, just like DNA.

A Nucleotide in RNA is still made of

3 important things:
1. 6-Carbon Sugar - Ribose
(instead of Deoxyribose)
2. Phosphate
3. Nitrogen base
there are 4 nitrogen bases in RNA,
A,G,C, and U that pair together)

AU C G
Ribonucleic Acid
RNA is SINGLE
STRANDED and does not
have to stay in the nucleus!

RNA is not found in

chromosomes because
it does not carry the
genetic code, however
it can read the DNA
code and take the
information out of
the nucleus.

RNA’s main job is to

build proteins!
 Three Main Types of
RNA
1. Messenger RNA (mRNA) - Carries copies of
instructions for the assembly of amino acids
into proteins from DNA to the rest of the cell
(serve as “messenger”)
 Three Main Types of
RNA
2.Ribosomal RNA (rRNA) – Makes up the major
part of ribosomes, which is where proteins are
made.

Ribosomal
RNA
 Three Main Types of
RNA
2.Ribosomal RNA (rRNA)
 Three Main Types of
RNA
2.Ribosomal RNA (rRNA)

• In 2009 Nobel Prize in

Chemistry awards V.
Ramakrishnan, Thomas A. Steitz
and Ada E. Yonath for having
showed what the ribosome
looks like and how it functions
at the atomic level.
 Three Main Types of
RNA
2.Ribosomal RNA (rRNA)

70S

S= Svedberg
Three Main Types of
RNA
 Three Main Types of
RNA
3. Transfer RNA (tRNA) - Transfers amino
acids to ribosomes during protein synthesis
3
A
C
C
A 5
C G
G C
C G
U G
U A
A U
UC AU
* CA C AG UA AG *
G * CU C *
C * GUGU * C GA G G
* UC * AGG
*GAG C
G C Hydrogen
U A bonds
* G
A
A* C
* U
A G
A
Anticodon
 2 Steps to Make a Protein
1. Transcription
• DNA → RNA

2. Translation
• RNA → Protein (Chain of amino acids)
• Transcription
o Initiation
o Elongation
o Termination

An overview of transcription. DNA binding at the promoter leads to

initiation of transcription by the polymerase holoenzyme, followed by
elongation and termination.
 Initiation
Where the transcription start?

• Transcription starts at promoters on the DNA

template. Promoter : A site in a DNA molecule at
which RNA polymerase and transcription factors
bind to initiate transcription of mRNA
 Initiation
RNA polymerase

Holoenzyme= 2   .

core enzym= RNA

polymerase without this
subunit (2   )
 Initiation
RNA polymerase
 Initiation
RNA polymerase
Initiation
 Initiation
DNA-Dependent Synthesis of RNA
 Like replication, transcription has initiation,
elongation, and termination phases.
 Transcription differs from replication in that it
does not require a primer and, generally,
involves only limited segments of a DNA
molecule.
 Additionally, within transcribed segments only
one DNA strand serves as a template for a
particular RNA molecule.
 This process is repeated many times as the
enzyme moves unidirectionally along the DNA
template.
Elongation
 Elongation
A segment of DNA has one strand
with the following sequence of bases:
AGC GCA TAG CAA
The complimentary strand of RNA
would be

a. UCG CGU AUC GUU

b. TCG CGT ATC GTT
c. AGC GCA UAG CAA
d. CTA TAC GCT ACC
o During elongation, the polymerase attempts to ensure
the accuracy of transcription by pyrophosphorolysis, in
which the catalytic reaction runs in reverse whenever
the polymerase stalls along the DNA.

o This process, known as kinetic proofreading, works

because the polymerase tends to stall after
incorporating a mismatched base into the growing RNA
chain, thus enabling pyrophosphorolysis to remove the
incorrect base.

o Pyrophosphorolysis is also used in the proofreading

that occurs during DNA synthesis.
• Proofreading by RNA polymerase. (a) In kinetic proofreading, the
polymerase stalls after incorporating a mismatched base into the
growing RNA chain, enabling pyrophosphorolysis to remove the
incorrect base. (b) In nucleolytic proofreading, the polymerase
backtracks on the DNA, melting several nucleotides of the RNA
(i.e., breaking the DNA-RNA base pairs), then an intrinsic nuclease
removes the section of melted RNA.
 Termination

o At this point, the polymerase releases the finished transcript

and dissociates from the template.

o E. coli DNA has at least two classes of such termination

sequences, one class that relies primarily on structures that
form in the RNA transcript and another that requires an
accessory protein factor called rho (ρ).

o Most ρ-independent termination sequences have two

distinguishing features.
Termination of transcription.
(a) In ρ-independent termination, an mRNA sequence forms a hairpin,
followed by Us’ residues, stalling the polymerase and separating it from the
mRNA.
(b) RNAs that include a rut site (purple) recruit the ρ helicase, which migrates
in the 5′→3′ direction along the mRNA and separates it from the polymerase.
 Differences
Between
Transcription In
Prokaryotes
and Eukaryotes
 Transcription And
Translation In Prokaryotes
5’ 3’

3’ 5’
RNA
Pol.

Ribosome

mRNA Ribosome
5’
 Eukaryotic
Transcription
Nuclear
Cytoplasm
pores
DNA

Transcription
RNA
RNA
Processing
mRNAG AAAAAA G AAAAAA

Export
Nucleus
 A “Simple” Eukaryotic
Gene
Transcription
3’ Untranslated Region
Start Site 5’ Untranslated Region
Introns

5’ Exon 1Int. 1 Exon 2 Int. 2Exon 3 3’

Promoter/ Terminator
Control Region Exons Sequence
RNA Transcript

5’ Exon 1Int. 1 Exon 2 Int. 2Exon 3 3’

 Processing Eukaryotic
mRNA
5’ Untranslated Region 3’ Untranslated Region

Protein Coding Region

5’5’ G 1Int. 1
ExonExon 1 Exon Exon 3 AAAAA3’
Int. 2
Exon 22Exon 3 3’

5’ Cap .1 .2 3’ Poly A Tail

t t
In In
 RNA processing achieves three
things:
1 Removal of introns (RNA splicing)
2 Addition of a 5’ cap
3 Addition of a 3’ tail (Poly A)
 This
signals the mRNA is ready to
move out of the nucleus and may
 Translation
involves “decoding” a messenger RNA
(mRNA) and using its information to build
a polypeptide, or chain of amino acids.
 The genetic code
Second mRNA base
U C A G
UUU UCU UAU UGU U
Phe Tyr Cys
UUC UCC UAC UGC C
U Ser
UUA UCA UAA Stop UGA Stop A
Leu
UUG UCG UAGStop UGG Trp G

Third mRNA base (3 end)

First mRNA base (5 end)

CUU CCU CAU CGU U

His
CUC CCC CAC CGC C
C Leu Pro Arg
CUA CCA CAA CGA A
Gln
CUG CCG CAG CGG G

AUU ACU AAU AGU U

Asn Ser C
AUC lle ACC AAC AGC
A Thr
AUA ACA AAA AGA A
Met or Lys Arg G
AUG ACG AAG AGG
start

GUU GCU GAU GGU U

Asp
GUC GCC GAC GGC C
G Val Ala Gly
GUA GCA GAA Glu GGA A
GUG GCG GAG GGG G
 Ribosomes
TRANSCRIPTION DNA

mRNA

TRANSLATION
Ribosome
Growing
Polypeptide polypeptide

tRNA
molecules Large
subunit
E
P A

Small
RNA + protein. subunit

We now think that RNA5

catalyzes the reactions. mRNA 3
 EPA model of a
ribosome
P site (Peptide site)
A site (Acceptor site)

E site (Exit site)

Large
subunit
E P A

mRNA
binding site
Small
subunit
New amino acids are added to the
carboxyl end of the polypeptide

Amino end Growing polypeptide

Next amino acid

to be added to
polypeptide chain

tRNA

mRNA 3

Codons
5
 The initiation of
translation Large
ribosomal
P site
3 U
A C 5 subunit
et et
M 5 A U G 3 M

Initiator tRNA
GTP GDP
E A
mRNA
5 3 5 3
Start codon
Small
mRNA binding site ribosomal Translation initiation complex
subunit
1 Small subunit binds just upstream 2 The large subunit can now
of start bind to make the active
ribosome
Special initiator methionine binds
to the start codon (AUG)
 TRANSCRIPTION DNA
Elongation
Amino end
1
tRNA with
of polypeptide
TRANSLATION
mRNA
Ribosome appropriate anticodon
Polypeptide
binds at A site
E
mRNA 3
Ribosome ready for P A
sitesite
next aminoacyl tRNA 5
2 GTP

2 GDP

E E

P A P A

2
GDP
3 GTP

Peptide bond
Translocaton. Everything E formed. Growing
moves left one notch. chain is now at A
Polypeptide is now at P. P A
Empty tRNA leaves the E
site.
 Termination
Release
factor
Free
polypeptide

5
3 3
3
5 5
Stop codon
(UAG, UAA, or UGA)

1 2 3
Stop codon has no Release factor The ribosome
matching tRNA binds and separates from
separates the mRNA
polypeptide from
the tRNA
 Antibiotics
1. Tetracycline: Blocks attachment of the Amino
Acid-tRNA to the A site in the ribosome

2. Erythromycin: Blocks the movement of the

ribosome down the mRNA

3. Streptomycin: Blocks formation of the

initiation complex and causes miscoding

4. Rifamycin: Blocks transcription by blocking

binding of RNA polymerase