Translation
Objectives
At the end of this
lecture, students will be able to –
• Describe the steps involved in the process of translation
• Explain post translational modifications
Content
• Translation process
• Post translational modifications
Translation
• The processed RNA molecule take part in protein synthesis
with the help of ribosomes.
• All the three types of RNAs are involved protein synthesis
• Three steps in protein synthesis
1. Initiation of the protein synthesis
2. Elongation of the protein synthesis
3. Chain termination
Initiation
in prokaryotes
• The essential factors required for initiation are
initiation factors, ribosome, mRNA, GTP, aminoacyl tRNA
• For initiation requires initiation factors like ІF 1, IF2,
IF3
• IF1 facilitates the attachment of the 30s subunit to the
mRNA and prevent the aa-tRNA from
entering the wrong site on the ribosome
• IF2 is a GTP binding protein required for attachment of
first aminoacyl- tRNA
• IF3 may prevent the large subunit 50s from joining prematurely
to 30s subunit
Binding the first
aa-tRNA into ribosomes:
N- formyl methionine
is the starting material . Aminoacylated initiator t RNA forms the
preinitiation complex by binding to the AUG codon of the mRNA and IF2
initiation factor is released
Assembling the
complete initiation complex:
Once the initiator tRNA is bound to the AUG codon, the large
subunit joins the complex and GTP bound to the IF2 is hydrolyzed and released
Elongation
• At the end of initiation sequence the 70s ribosome
possesses the fmet-tRNA in the p site, where as the A site is free to receive
the next aminoacyl- tRNA according to the codon on mRNA
• The elongation factor
(EF) is a
soluble protein which
is required for elongation of peptides
• two types of EF-T
and EF-G
• EF-T is associated
with tranferase activity( EF-Tu and EF-Ts)
• EF-G is involved in traslocation of mRNA
There are 3 steps in elongation
1. Binding of aminoacyl tRNA to A site
2. Peptide bond formation
3. Translocation.
Binding of aminoacyl
tRNA to A site:
• 70s initiation complex contains fmet tRNA in the P site
and A site is free
• Before the
second aminoacyl tRNA is placed in the A site it must combine
with the protein EF linked to the GTP
this EF is called as EF-Tu Once the proper t-RNA Tu GTP is bound to the mRNA
codon , GTP is hydrolysed and Tu- GDP complex released , leaving the
aa- tRNA in the A site
Peptide bond
formation
• Enzyme peptidyl transferase catalyzes the formation of
peptide bond
• The net result of peptide bond formation is attachment of
growing peptide chain to tRNA in A site
Traslocation
• As peptide bond occurs, the ribosomes moves to the next
codon of mRNA this is called translocation
• Involves the movement of growing peptide chain from A site
to P site
• Requires EF-G and GTP.
GTP gets hydrolyzed and supplies energy to move mRNA
Termination
• After several cycles of elongation, incorporating the
amino acid and the formation of specific protein / polypeptide molecule, one of
the stop codon (UAA, UGA, and UAG) terminates the growing peptide
• Termination codon occupies ribosomal A site, the release
factor (RF1, RF2, RF3) recognizes the stop signal
• RF1 is active with UAA and UAG codons RF2 is active with
UAA and UGA. RF3 activates the RF1 and RF2 i.e it is called stimulatory factor
• Ribosome binds either with RF1 or RF2 activates the
peptidyl transferase which hydrolyze the bond joining the peptide to the tRNA
at the P site this result in release of the peptide chain
Initiation
in eukaryotes
• It requires 10 initiation factors out of which eIF1, eIF2,
eIF3 binds to the 40s subunit, which prepares the subunit for binding to mRNA
• The initiator tRNA (methionine) also binds to 40s subunit prior
to its interaction with mRNA
• tRNA enters in association with eIF2 GTP
• This complex is called 43s preinitiation complex
• eIF4E binds to the 5ꞌ
cap of eukaryotic mRNA
• eIF4A moves along the 5ꞌ
end of the message removing any double stranded region that would interfere
with the movement of 43s complex
• Once the 43s complex binds to the 5ꞌ end of the mRNA, the complex scans along the
message until it reaches a recognizable sequence of nucleotide (typically 5ꞌ CCACCAUGC- 3ꞌ) that contains AUG initiator
codon
• Once it reaches the AUG eIF2 – GTP is hydrolyzed and
released the large subunit 60s joins the complex to complete intiation
Post – translational modification
The protein synthesized in translation are as such not
functional
Many changes takes place in polypeptides after the protein
synthesis
1. Protein folding
2. Proteolytic degradation
3. Intein splicing
Protein folding:
the three dimensional conformation of proteins is important for their
biological functions
Chaperons are
heat shock proteins facilitate and favour the interaction on polypeptides
surface to give the specific conformation of protein
• Chaperons reversibly bind to the hydrophobic region of unfolded
proteins and Folding intermediates which helps the protein to attain compact
and biologically active conformation
Proteolytic
degradation:
• Many proteins are synthesized as precursors which are
bigger than the functional protein
• Some portion of precursor molecules are Removed by
proteolysis to liberate active proteins
• This process is called as Trimming
Intein splicing:
• Inteins are intervening sequence in certain proteins
• These are comparable to introns in mRNA
• Inteins have to be removed and exteins ligated in
appropriate order for protein to become active
Summary
• Translation involves the conversion of mRNA into proteins
• Steps involved are initiation, elongation and translation
• The essential factors required for initiation are
initiation factors, ribosome, mRNA, GTP, aminoacyl tRNA
• Changes takes place in polypeptides after the protein
synthesis -Protein folding, proteolytic degradation, intein splicing