Chapter 24

Translation

Structure is evolutionarily conserved, yet overall size and
components vary
24.1 Introduction
llgure 24.1

Ribosomes are ribonucleoprotein particles.
24.1 Introduction
llgure 24.2

An aminoacyl-tRNA brings an amino acid
to ribosome and enters the A site.
Peptidyl-tRNA is bound in the P site.
Deacylated tRNA exits via the E site (in
bacteria eukaryotic ribosome does not
have E site).
An amino acid is added to the polypeptide
chain by transferring the polypeptide from
peptidyl-tRNA in the P site to aminoacyl-
tRNA in the A site.
A & P sites occupy both large and small
subunits of ribosome.
24.2. Translation Occurs by
Initiation, Elongation, and
Termination
llgure 24.4
Initiation: all the processes
before forming the peptide bond
between the first two amino
acids of the protein (e.g.,
ribosome binding to mRNA) !
slow and rate-limiting step
Elongation: fastest step in
translation. An amino acid is
added to the polypeptide chain
by transferring the polypeptide
from peptidyl-tRNA in the P site
to aminoacyl-tRNA in the A site
(translocation)
Termination: release of
polypeptide and ribosome
dissociation
24.2. Translation Occurs by Initiation,
Elongation, and Termination
llgure 24.08: 1ranslauon falls lnLo Lhree sLages.
Ribosome and 30S/50S
subunits are in
equilibrium.
30S subunit binds to
mRNA first and 50S is
recruited.
Ribosome assembly on
mRNA requires initiation
factors (IFs)
Ribosome is released
as free ribosome or
ribosomal subunits.
24.4. Initiation in Bacteria Needs 30S
Subunits and Accessory Factors
llgure 24.10
Ribosome-binding site (rbs):
a short sequence of bases
that precedes the start
codon.
Shine-Dalgarno sequence
(AGGAGG): ~10 base
upstream of AUG,
complementary to 3 end of
16S rRNA.
Also read section 24.6.

24.4. Initiation in Bacteria
Needs 30S Subunits and
Accessory Factors
llgure 24.13
24.4. Initiation in Bacteria
Needs 30S Subunits and
Accessory Factors (=
Initiation Factors, IFs)
Initiation factors (IF-1, -2,
and -3), which bind to 30S
subunits, are required for
translation initiation.
A 30S subunit carrying
initiation factors binds to an
initiation site on mRNA to
form an initiation complex.
IF-2 binds to initiator tRNA
and brings it to P site
element in 30S subunit/
mRNA complex
IF-1 binds to near A site and
prevents aminoacyl-tRNA
from entering.
llgure 24.11
IF-3 controls
equilibrium between
ribosomal states
IF-3 enables 30S to
bind to mRNA
IF-3 checks the
accuracy of recognition
of the first aminoacyl-
tRNA
Must be released to
allow 50S subunits to
join the 30S/mRNA.
llgure 24.11
24.4. Initiation in
Bacteria Needs 30S
Subunits and
Accessory Factors
Translation starts with a methionine
amino acid usually coded by AUG
(GUG and UUG can be used in
bacteria).
In bacteria, mitochondria, and
chloroplast different methionine
tRNAs are involved in initiation (N-
formyl-methionyl-tRNA; fMet-tRNA
f
)
and elongation (Met-tRNA
m
).
The initiator tRNA has unique
structural features:
No base pairing of the first base at 5
end
Three G-C pairs in the stem preceding
the anticodon loop
Formylated amino acid
24.5 A Special Initiator tRNA Starts the Polypeptide Chain
llgure 24.14
!"
$"
%&'()*)&
Eukaryotic 40S ribosomal
subunits bind to the 5! cap of
mRNA and scan the mRNA until
they reach the initiation site.
A eukaryotic initiation site
consists of a ten-nucleotide
sequence that includes an AUG
codon (Kozak consensus
sequence).
Internal Ribosome Entry Site
(IRES): ribosome binds directly
to mRNA structure containing
AUG start codon
24.7 Small Subunits Scan
for Initiation Sites on
Eukaryotic mRNA
llgure 24.17
Initiation factors are required for all
stages of initiation, including:
binding the initiator tRNA
i
(eIF2)

40S subunit attachment to
mRNA
movement along the mRNA
joining of the 60S subunit
eIF2 and eIF3 bind the initiator
Met-tRNAi and GTP.
The complex binds to the 40S
subunit before it associates
with mRNA.
eIF4G links 5 and 3 end of
mRNA.
24.7 Small Subunits Scan
for Initiation Sites on
Eukaryotic mRNA
llgure 24.18:
24.8. Elongation Factor Tu (EF-Tu) Loads Aminoacyl-
tRNA into the A Site
EF-Tu is a monomeric G protein
whose active form (bound to
GTP) binds aminoacyl-tRNA.
The EF-Tu-GTP-aminoacyl-tRNA
complex binds to the ribosome A
site (P site is occupied).
Once codon-anticodon base-
pairing is complete, Tu
hydrolyzes GTP (releasing Tu-
GDP) and tRNA occupies A site
in 50S; Tu-GDP is converted to
Tu-GTP by EF-Ts.
In eukaryotes, eEF1" & eEF1#$
are counterparts of EF-Tu & EF-
Ts

llgure 24.19: Lf-1u-C1 places amlnoacyl-L8nA on
Lhe rlbosome and Lhen ls released as Ll-1u-Cu.
The 50S subunit has peptidyl
transferase activity.
The nascent polypeptide chain is
transferred from peptidyl-tRNA
in the P site to aminoacyl-
tRNA in the A site.
Peptide bond synthesis
generates deacylated tRNA in
the P site and peptidyl-tRNA in
the A site.
24.9 The Polypeptide Chain Is Transferred to
Aminoacyl-tRNA
llgure 24.20
+ ,-./
0 ,-./
1 ,-./
24.10 Translocation Moves
the Ribosome
Ribosomal translocation
moves the mRNA through
the ribosome by three bases.
Translocation moves
deacylated tRNA into the E
site and peptidyl-tRNA into
the P site, and empties the A
site.
llgure 24.22
24.10 Translocation Moves the
Ribosome
The hybrid state model
proposes that translocation
occurs in two stages, in which
the 50S moves relative to the
30S and then the 30S moves
along mRNA to restore the
original conformation.
llgure 24.23: Models for
Lranslocauon lnvolve Lwo sLages.
24.11 Elongation Factors Bind
Alternately to the Ribosome
Translocation requires EF-G
(eEF2 in eukaryotes), whose
structure resembles the
aminoacyl-tRNA-EF-Tu-GTP
complex.
Binding of EF-Tu and EF-G to the
ribosome is mutually exclusive.
Translocation requires GTP
hydrolysis, which triggers a
change in EF-G, which in turn
triggers a change in ribosome
structure.
llgure 24.24
24.11 Elongation Factors Bind Alternately to the
Ribosome
llgure 24.23
23./45-&6,
.780
24.12 Three Codons Terminate Translation and Are
Recognized by Protein Factors
Three stop codons UAG (amber), UAA (ochre) and
UGA (opal) terminate translation.
In bacteria they are used most often with relative
frequencies UAA>UGA>UAG.
Termination codons are recognized by release factors
(RFs), not by aminoacyl-tRNAs.
RF1, RF2 and RF3 in bacteria; eRF1 and eRF3 in
eukaryotes.
24.12 Three Codons
Terminate Translation and Are
Recognized by Protein Factors
The structures of the class 1
release factors (RF1 and RF2 in
E. coli ) resemble aminoacyl-
tRNA%EF-Tu and EF-G.
RF1 recognizes UAA and UAG;
RF2 recognizes UGA and UAA.
These factors bind to A site only
when P site is occupied.
Class 2 RF (RF3 in E. coli)
releases class I RFs from
ribosome.
llgure 24.26: Molecular mlmlcry enables
Lhe elongauon facLor 1u-L8nA complex,
Lhe Lranslocauon facLor Ll-C, and Lhe
release facLors 8l1/2-8l3 Lo blnd Lo Lhe
same rlbosomal slLe.
24.12 Three Codons
Terminate Translation and Are
Recognized by Protein Factors
Termination reaction releases
the polypeptide (protein) but
leaves a deacylated tRNA and
mRNA still associated with
ribosome.
Ribosome recycling factor (RRF)
along with EF-G dissociate
ribosome and uncharged tRNA.
IF3 binds to 30S and inhibits
reassociation.
llgure 24.28: 1he 8l LermlnaLes proLeln
synLhesls by releaslng Lhe proLeln chaln.
1he 88l releases Lhe lasL L8nA, and Ll-C
releases 88l, causlng Lhe rlbosome Lo
dlssoclaLe.
24.12 Three Codons Terminate Translation and Are
Recognized by Protein Factors
llgure 24.29: luncuonal homologles of prokaryouc and eukaryouc Lranslauon facLors.
16S rRNA plays an active role in the functions of the 30S
subunit.
It interacts directly with:
mRNA (i.e., rbs)
23S rRNA in the 50S subunit (ribosome assembly)
the anticodon regions of tRNAs in P site and A site
23S rRNA interacts with the CCA terminus of peptidyl-
tRNA in both P site and A site.
Peptidyl transferase activity resides exclusively in the 23S
rRNA but not proteins.
24.15 Two rRNAs Play Active Roles in Translation
24.16 Translation Can Be Regulated
Translation can be regulated by the 5! UTR of the mRNA.
Translation may be regulated by the abundance of various
tRNAs = codon usage.
A repressor protein can regulate translation by preventing
a ribosome from binding to an initiation codon.
llgure 24.40: A regulaLor proLeln may
block Lranslauon by blndlng Lo a slLe on
m8nA LhaL overlaps Lhe rlbosome-
blndlng slLe aL Lhe lnluauon codon.
24.16 Translation Can Be
Regulated
Translation initiation region
sequence context.
Accessibility of initiation codons
in a polycistronic mRNA can be
controlled by changes in the
structure of the mRNA that occur
as the result of translation of a
preceding gene.
microRNA-mediated translation
suppression.
llgure 24.42: Secondary
sLrucLure can conLrol lnluauon.