Translation in Detail

translation is a complex process in

which a ribosome uses the coded

information found in messenger RNA or

mRNA to build a functional protein when

you have completed this exercise you

should understand the structure and

function of the ribosome understand the

function of charged trnas understand the

three distinct steps of translation

initiation elongation and termination

and understand the functions of

translation factors in each of these

steps

the genetic information necessary to

build proteins is contained within an

organism's genome to create a protein

genes are first transcribed to create

messenger RNA the MRNA is then read by a

large structure called the ribosome

which links amino acids together to form

the protein encoded by the MRNA

the ribosome cannot accomplish this feat

of aided however besides the MRNA and a

ribosome translation also requires amino

acid carriers called transfer rnas and a

large number of initiation elongation

and release factors finally ribosomes

cannot be reused in a Cell without the

action of a recycling Factor

the ribosome is a large macromolecular

structure that performs protein

synthesis in cells the ribosome links

together the appropriate amino acids to

form the polypeptide chain encoded by

the gene

the ribosome consists of two subunits

the small subunit and the large subunit

each consisting of both ribosomal RNA

and protein

in contrast with many enzymes you have

studied over half of the ribosome is

composed of RNA with protein making up a

smaller portion of the mass of this

enzyme ribosomes are similar in

structure in prokaryotes and eukaryotes

although the overall size of the

ribosome is smaller in prokaryotes

ribosomal subunits are named by their

svedberg coefficients a measure of their

sedimentation rate when subjected to

centrifugal force

ribosomal subunits assemble into a full

ribosome on the MRNA disassemble when

the protein has been synthesized and

assemble once again to start another

round of protein synthesis this process

of repeated assembly and disassembly of

ribosomal subunits is known as the

ribosome cycle

the ribosome reads the MRNA three bases

at a time note that while spaces have

been added to Aid in visualization these

triplet codons are immediately adjacent

to one another on the MRNA the triplet

codons are matched within the ribosome

to specialized RNA molecules that carry

amino acids these amino acid carrying

rnas are called transfer rnas or trnas

for they are adapter molecules that

transfer genetic information to amino

acid sequence

trnas contain anticodons that are

complementary to and thus bind the

codons on the MRNA

the ribosome is uniquely designed to

bring mRNA and trnas together the MRNA

is threaded through the small subunit

while the large and small subunits of

the ribosome put together to make TRNA

binding pockets

the ribosome contains three binding

sites for holding TRNA the a site

accepts tRNA molecules charged with

single amino acids the P site contains

the TRNA linked to the growing peptide

chain and the e or exit site is occupied

by a TRNA molecule about to be released

the growing protein exits the ribosome

through a tunnel in the large subunit

for translation to begin the ribosome

needs to be seated properly over the

start codon of the MRNA with a Charged

TRNA in the P site this process of

initiation which differs significantly

in prokaryotes and eukaryotes is aided

by a number of translation initiation

factors

first we will describe prokaryotic

translation initiation which requires

three initiation factors if1 if2 and if3

each binds the 30s small ribosomal

subunit

initiation Factor 3 binds the 30s small

subunit and prevents association with

the 50s large subunit this keeps the 30s

small subunit open for mRNA binding next

initiation Factor 1 nears the a site and

initiation Factor 2 binds both the 30s

subunit and if1 in association with a

GTP molecule

next both the MRNA and initiator TRNA

can assemble onto the complex initiation

Factor 1 blocks trnas from improperly

entering the a site and if2 brings the

initiator TRNA charged with s methionine

to the P site

the ribosome binds to a sequence on the

MRNA called the shine delgarno sequence

through a complementary sequence on the

16s RNA of the small subunit the

messenger and transfer rnas combine the

30s subunit in either order

the small subunit of the ribosome

aligned with the MRNA start codon is

called the 30s initiation complex

The Binding of the charged ethnet TRNA

to the small subunit initiates a

sequence of events to form the 7ds

initiation complex once the fmat TRNA

has bound the small subunit if-3 falls

off and is replaced by the large subunit

which completes the ribosome large

subunit binding stimulates the gtpase

activity of if2 specifically the factor

binding Center on the large subunit

stimulates if2's gtpace activity

because the GDP if2 complex has a low

affinity for the ribosome if2 and GDP

and then if1 are all released this is

the 7ds initiation complex protein

synthesis can now begin

in eukaryotic translation initiation is

more complex and uses at least eight

initiation factors compared to three in

prokaryotes to begin the small ribosomal

subunit binds to the five Prime end of

the MRNA called a five Prime cap and

then scans along the MRNA to find the

start codon

translation factors eif3 and eif 1A bind

the 40s small ribosomal subunit to

prevent the association of the large 60s

subunit before the MRNA can bind

next eif-5b recruits an eif-2 GTP met

initiator TRNA complex to the subunit

where eif-2 and eif-5b place the

initiator TRNA in the P site this forms

the 43s pre-initiation complex note that

this pre-initiation complex does not

contain messenger RNA

meanwhile

eif4f which consists of the three

subunits eif4a eif 4E and eif 4G binds

to the five Prime end of the eukaryotic

mRNA the 4E subunit binds the five Prime

cap A specialized structure at the very

tip of the MRNA while the 4A and four G

subunits bind further down eif 4A is

needed for the MRNA unwinding that will

allow the ribosome to find the start

codon of the MRNA

next eif-4b binds to the eif4f complex

and activates a helicase activity this

unwinds any secondary structure in the

MRNA the 43s pre-initiation complex can

now bind to the MRNA through the

interaction of eif3 and eif4f

once bound the 40s subunit scans along

the message aided by the eif-4f helicase

activity until the anticodon of the

initiator Met TRNA recognizes the AUG

start codon on the MRNA

eif-2 and eif3 are released from the

complex and the 60s large subunit

completes the ribosome the remaining

initiation factors are released after

large subunit binding stimulates the

eif-5b gtph activity as with prokaryotic

if2 the gtpace activity of eif-5b is

activated by contact with the factor

binding Center on the large ribosomal

subunit the ads initiation complex is

now ready to begin protein synthesis

after an initiation complex is formed

elongation occurs during elongation the

ribosome synthesizes proteins by linking

together amino acids three steps are

needed to add each amino acid a Charged

TRNA is placed in the a site of the

ribosome a peptide bond is formed and

the TRNA with the growing protein chain

is moved from the a site to the P site

in prokaryotes this requires two

translation elongation factors eftu and

EFG a third elongation factor efts is

needed to recycle eftu elongation is

very similar in eukaryotes and only the

prokaryotic process will be described

here

when the triplet anticodon of the

correct TRNA is base paired with the

MRNA in the ribosome a site the base

pairing is strengthened by additional

hydrogen bonding with the ribosome

incorrectly bound trnas do not base

pairs strongly with the ribosome and are

released before they can be added to the

growing polypeptide chain correctly base

paired trnas have additional hydrogen

bonds that help the TRNA remain in the a

site during accommodation the process

whereby the TRNA rotates into position

for peptide bond formation

first a Charged TRNA is bound by eftu

and is brought to the ribosome a site as

an eftu GTP amino acid TRNA complex the

TRNA anticodon binds the MRNA codon and

eftu hydrolyzes its GTP and the

resulting eftu GDP is released from the

TRNA

next a peptide bond is formed between

the amino acids this reaction transfers

the amino acid from the TRNA in the P

site onto the amino acyl TRNA in the a

site

to prepare for addition of the next

amino acid the ribosome undergoes

translocation a process in which the

ribosome moves along the MRNA to the

next codon translocation is facilitated

by elongation Factor EFG the TRNA with

the growing amino acid chain attached

moves from the a site into the P site

and the spent TRNA in the P site moves

into the E site

note that elongation factors eftu and

EFG both end up as complexes with GDP

after one round of elongation both

factors must exchange their GDP for GTP

before they can be used again for eftu

this exchange is facilitated by

elongation Factor efts

for EFG the GDP simply falls off and is

quickly replaced by a GTP

process continues at

on the corresponding amino acid TRNA

binds in the a site the growing peptide

chain attaches to the amino acid on the

TRNA in the a site and the ribosome

translocates moving the growing protein

chain and TRNA into the P site and

freeing up the a site for another round

of amino acid addition

termination of translation is engaged

when a stop codon on the MRNA enters the

a site of the ribosome and is performed

by release factors that recognize the

stop codon

the mechanism of termination is similar

in prokaryotes and eukaryotes and

involves two kinds of release factors

called class 1 release factors and Class

2 release factors

class 1 release factors enter the

ribosome a site and bind the stop codon

using a peptide anticodon prokaryotes

have two class one release factors rf1

and rf2 eukaryotes have one class one

release Factor erf1

the class 2 release factor in complex

with GDP also binds the large subunit in

prokaryotes rf3 is the class 2 release

factor in eukaryotes erf3 is the class 2

release Factor after the class 1 and

Class 2 release factors are both bound a

glycine glycine glutamine or ggq Motif

on the class 1 release Factor triggers

hydrolysis of the protein from the TRNA

in the P site

release of the protein from the ribosome

triggers the recycling of the release

factors first GTP is exchanged for GDP

on the class II release Factor this

causes the class 2 release factor to

push the class 1 release Factor off the

ribosome

the release of the class 1 release

Factor allows rf3 to interact with the

factor binding Center which stimulates

rf3's gtpace activity GTP is then

hydrolyzed on the class 2 release Factor

the class 2 release Factor GDP complex

then releases from the used ribosome

at this point the ribosome still

contains two de-acetylated trnas and is

also still bound to the MRNA the process

of cleaning out the ribosome and

separating the subunits is called

ribosome recycling although not as well

understood in eukaryotes in prokaryotes

this process involves a ribosome

recycling Factor plus initiation factor

if3 and elongation Factor EFG

the ribosome recycling Factor rrf binds

to the a site of the used ribosome and

recruits EFG to the complex the rrf acts

like an a site TRNA during the

translocation process

EFG can thus push on the rrf in the same

way as it would on a TRNA this moves the

rrf into the P site and releases the two

spent trnas next both the rrf and EFG

uncoupled from the ribosome the MRNA is

released possibly with the aid of if3

if3 binds the small subunit to prepare

it for another round of protein

synthesis