TRANSLATIO

N
Translation
• Process wherein polypeptide chains are
synthesized
• Uses mRNA as template which determines the
order in which different amino acid residues are
to be joined
Translation: Prokaryotes vs. Eukaryotes
THE TRANSLATION
MACHINERY
mRNA
• Template for translation
• Codon
• Three adjacent ribonucleotides along the
mRNA which correspond to a specific
amino acid
The Genetic Code
• The correspondence between codons and
amino acids
The Genetic Code: Properties
• Universal
(almost)
Codon Universal code Unusual code Occurence

UGA stop Trp Mycoplasma, mitochondria

UGA stop Cys Euplotes

UAA, UAG stop Gln Various protozoa

CUG Leu Thr Yeast mitochondria

CUG Leu Ser Candida

The Genetic Code: Properties
• Degenerate
• Some amino acids are coded by more than
one codon
• e.g. Phe is coded by UUU and UUC
• Contiguous
• Codons do not overlap and have no
spacers in between
• Unambiguous
• Each codon specifies only one amino acid
Prokaryotic mRNA
• Contains the Shine-Dalgarno Sequence
• Found near the 5’-end of the mRNA
• Sequence responsible for proper positioning
of the mRNA in the ribosome
• Complementary to a portion of the 16S rRNA
in the 30S (small) ribosomal unit
• Consensus sequence: 5’-AGGAGGU-3’
tRNA
• Carries the amino
acid to the
translation
machinery
• Contains the
anticodon that is
complementary to
the codon in the
mRNA
• Has a cloverleaf
structure
Parts of tRNA
• Acceptor stem
• the 5’-CCA-3’
terminal that binds
the amino acid
through an
ester bond
• Anticodon arm
• contains the
anticodon that is
complementary to
the codon it
recognizes
in the mRNA
Parts of tRNA
• D or DHU arm
• TΨCG arm
• contribute
important
interactions for
the overall
folding of tRNA
molecules
• Rich in unusual
nitrogenous
bases
Tertiary Structure of tRNA
• Inverted L-shaped structure
• Formed by 9 H-bonds mainly between the
unusual bases in the variable regions
The Genetic Code

• 64 codons – 3 stop codons = 61 tRNAs

needed
The Wobble Hypothesis
• Rules of base pairing
are relaxed in the third
position of the codon
• Reduces the
number of tRNAs
needed by the cell
• Enables one tRNA
to read two or
three codons
The Wobble Hypothesis
Ribosomes
• Composed of:
• Large subunit
• contains the peptidyl transferase center -
catalyzes peptide bond formation
• factor-binding center
• Small subunit
• contains the decoding center: narrow
channels where the mRNA enters and exits
Ribosomes
• A site
• Aminoacyl
tRNA binding
site
• P site
• Peptidyl tRNA
binding site
• E site
• Exit site
Sedimentation Coef ficient
• a measure of the rate of sedimentation in an
ultracentrifuge of a molecule suspended in a
less dense solvent.
• measured in Svedberg units, S.
• S values are not additive.
Eukaryotic Ribosomes
• Large subunit – 60S
• 50 proteins + (28S + 5.8S + 5S)
rRNA
• Small subunit – 40S
• 33 proteins + 18S rRNA
Prokaryotic Ribosomes
• Large subunit – 50S
• 31 proteins + (23S + 5S)
rRNA
• Small subunit – 30S
• 21 proteins + 16S rRNA
PROKARYOTIC TRANSLATION
Amino Acid Activation

Alberts et al
2008
Amino Acid Activation
Amino acid + ATP →
amino acyl-AMP
+ PPiAmino acyl-AMP
• Catalyzed by: amino acyl-tRNA + tRNA
synthetase →
• Eg.amino
for the activation
acyl-tRNA of alanine
+ AMP
• Enzyme: alanine-tRNA synthetase
• Activated form: alanine-tRNAAla
Initiation Complex Formation
• Initiator tRNAifmet
• recognizes AUG (and GUG
or UUG by wobble)
• aminoacylated with met
• met is formylated in its
amino group to become
N-formylmethionine (fmet)
• f-met as temporary N-
terminus of the peptide
Initiation Complex Formation
1. IF1 promotes dissociation of ribosomal
subunits
2. IF3 binds to 30S subunit to prevent its re-
association with the 50S subunit
Initiation Complex Formation
3. mRNA binds to the 30S subunit
• Shine-Dalgarno sequence as ribosome
binding site
Initiation Complex Formation
4. IF2 (along with
GTP) brings the
initiator tRNA fmet i
to the 30S subunit.
Initiation Complex Formation
5. IF1 bound to the
initiation complex
• induces
conformational
change
• allows the 50S
subunit to attach
using energy from
GTP hydrolysis.
• IF’s are then
released.
Elongation of Polypeptide
1. Binding of the next charged amino acid
• A-site as entry point
• Facilitated by EF-Tu and requires hydrolysis of
GTP
• EF-Ts recharges EF-Tu with GTP
2. Peptide bond formation
• Polypeptide from the tRNA in the P-site forms
a peptide bond with the amino acid attached
to the tRNA in the A-site
• Catalyzed by peptidyl transferase
Elongation of Polypeptide

EF-
Tu

Peptide bond
formation
catalyzed by
peptidyl transferase
Elongation of Polypeptide
3. Translocation
• Ribosome slides through the mRNA to read
the next codon
• Promoted by another elongation factor EF-G
and requires GTP hydrolysis
• Places empty tRNA in the E-site
• tRNA with growing polypeptide chain now on
the P-site
Elongation of Polypeptide

EF-
G
Termination of Translation
1. Recognition of stop codons in the A-site
• UAG, UGA, UAA
2. Binding of release factor (RF) in the A-site
• RF1 recognizes UAG and UAA;
• RF2 recognizes UGA and UAA;
• RF3 assists RF1 or RF2
• Hydrolyzes the polypeptide chain from the
tRNA
3. Dissociation of the translation machinery
Termination of Translation
Transcription and Translation
EUKARYOTIC TRANSLATION
Initiation Factors
Prokaryotic Eukaryotic Function
IF1, IF3 eIF3, eIF4C, eIF6 • Binding to ribosome
subunits
IF2 eIF4B, eIF4F eIF2, • Binding to mRNA
eIF2B eIF5 • Initiator tRNA delivery
• Displacement of other
factors
Translation Initiation
• Prokaryotes
• translation initiation complex is built directly
on start codon
• facilitated by the Shine-Dalgarno sequence
• Eukaryotes
• ribosomes indirectly locate start codon
• there is no ribosome-binding site in
eukaryotic mRNA
• involves a scanning mechanism starting at
the 5’
cap
Translation Initiation
• Scanning
Mechanis
m
Translation Initiation
• Scanning
Mechanism
Translation Elongation
• Elongation Factors:
• eEF-1α (= EF-Tu)
• eEF-1βγ (= EF-
Ts)
• eEF-2 (= EF-G)

• no E-site in
ribosome
Protein Targeting in Eukaryotes
• to target proteins into their respective
location within or outside of the cell before
they become biologically active
• Involves:
• signal peptide
• in the N-terminal of the nascent protein
• signal recognition particle (SRP)
• recognizes signal peptide
Events in Protein Targeting

• Glycosylation pattern (step 6) control the

f inal location of the protein.