CHAPTER 40

The Mechanism of Protein Synthesis

Problems: 2,3,6,7,9,13,14,15,18,19,20

• Initiation: Locating the start codon.

• Elongation: Reading the codons (5’→3’) and

synthesizing protein amino→carboxyl.

• Termination: Recognizing terminal codon

and releasing protein.

Prokaryotes and Eukaryotes

40:1 Decoding the Information in
mRNA

Exit Peptidyl Aminoacyl

Initiation

Start codon (also GUG, Val)

Shine-Delgarno Sequence

Start:
1. Pairing of 16S
rRNA with S-
D of mRNA (a
UTR).
2. Pairing of
initiator tRNA
with start
codon.
 Initiator tRNAf
Two different tRNA’s that
carry Met; tRNAf and tRNAm.

One Synthetase adds Met to

both tRNAf and tRNAm.

A specific transformylase only

formylates Met-tRNAf.
 Initiation
Initiation factors, IF1, IF2
and IF3.

IF1 and IF3 prevent

premature assembly of the
70S ribosome.

IF2 binds to fMet-tRNAf

and mRNA at AUG codon.

The fMet-tRNAf in the fully

assembled ribosome is in the
P-site.
 Initiation
Elongation Factor, EF-Tu
brings appropriate aa-tRNA fMet

to the A-site.

AUG

Requires GTP: EF-Tu-GTP

Protects aa-tRNA ester
bond.

GTP→GDP only when proper

codon is present and then
EF-Tu is released.
40:2 Peptidyl Tranferase and Peptide
Bond Formation---Elongation

Peptidyl tranferase
center on 23S rRNA
of 50S subunit.

Catalysis occurs via

mechanism of
proximity and
orientation.
 EF-Tu-GTP

EF-Ts GDP

GTP
-EF-Tu-GTP
EF-Tu-GDP
-EF-Tu-GDP
Peptide grows from the H3N+-end to –COOH-end
 Termination

Release Factors (RF) recognize the stop codons: RF1-UAA

or UAG and RF2-UAA or UGA.

RF recognizes stop codon and interacts with peptidyl

transferase promoting addition of H2O, rather than
aminoacyl-tRNA, to the growing peptide chain.

GTP hydrolysis after binding EF-G and binding of ribosome

release factor (RRF) cause release of ribosome.
40:3 Bacteria and Eukaryotes Differ
in Initiation of Protein Synthesis
Larger ribosome: 80S versus 70S.

Initiator tRNAi and Met-tRNAi is not

formylated; differs from Met-tRNAm.

Initiation involves eIF’s, CAP recognition

(eIF-4E), and movement to AUG start codon
(eIF-2).

Circular structure of eukaryotic mRNA

thought to facilitate rebinding of ribosomes
Important control
point. Mutation leads
to serious disease.
Elongation: EF1 and EF1 are counterparts
of EF-Tu and EF-Ts, and EF2 is equivalent of
bacterial EF-G translocase.

Termination: eRF1 is the release factor and is

the only release factor in eukaryotes. eIF-3
prevents premature ribosome reassembly in
absence of initiation factors.

Organization: In eukaryotes the translation

machinery is organized as a large complex with
the cytoskeleton.
40.4:
Antibiotic
Protein
Synthesis
inhibitors.

Diphtheria toxin
inhibits EF2 (the
eukaryotic
translocase).
 40:5 Secretory and Membrane
Proteins
Protein targeting or sorting: 1. Posttranslational delivery to nucleus,
chloroplast, mitochondria, and peroxisomes. 2. Secretory pathway to
ER, and then the Golgi, lysosomes, integral membrane proteins, etc.

Signal
recognition
particle

Packaged in transport vesicles

and transported to location via
exo- and endocytosis
 40:6 Protein Synthesis Regulation
Regulation by use of mRNA: Iron Response Elements (IRE)
Ferritin mRNA contains an IRE to
which an IRE binding protein (IRE-BP)
binds and blocks translation of the
mRNA.

High iron: IRE-BP binds iron and can

not bind ferritin mRNA. Ferritin is
produced to store iron.

Low iron: IRE-BP binds ferritin mRNA

and blocks ferritin production

Transferrin-receptor mRNA
contains an IRE to which an IRE
binding protein (IRE-BP) binds
under low iron that stabilizes the
mRNA, permitting translation and
production of the receptor.
 Regulation Through Small RNAs
RNA interference, RNAi: External source of dsRNA is cleaved into
21-nucleotide fragments by an RNase → ss siRNA → RISC which
locats siRNA onto complementary mRNA and promotes their
degradation.

MicroRNAs, miRNA: miRNA produced from endogenous encoded

RNA precursor this binds to complementary region of mRNA forming
a 21-nucleotide ds RNA region which promotes degradation of that
mRNA. Human genome has more that 700 miRNAs, 60% of genes
regulated by one or more miRNAs: cell differentiation, development,
disease such as cancer, etc.