TRANSLATION

The translation process is the rendering of the nitrogen bases sequence in the mRNA into the
sequence of amino acids in the protein. It is the process of synthesis of proteins, during which the
amino acids are ordered according to the triplet sequence in a given mRNA. The proteins are then
changed by post-translation modifications, first in the endoplasmic reticulum and the Golgi
apparatus. The correct primary sequence of the amino acids in the chain allows the creation of an
active secondary, tertiary, or quarterly structure, which is necessary for protein full biological
activity.

The protein synthesis in the cell takes place in the ribosomes, in the cytoplasm and endoplasmic
reticulum. First is necessary to build (assembly) a protein synthetic apparatus, which consists
of: mRNA, small and big subunit of ribosome, aminoacyl-tRNA synthetase, helping molecules
and amino acids.

The process of translation can be divided into the following stages:

1. The creation of an initiation complex;
2. The activation of an amino acid and its bond on the tRNA;
3. The lengthening of the peptide chain;
4. The termination of the synthesis of the polypeptide;
5. Post -translation modification of the protein chain.

The formation of the initiation complex

Here the mRNA binds to the small subunit of ribosome by the cap and 5´UTR, with the help of
regulatory proteins. The tRNA binding methionine amino acid (the one complementary to the
start codon) also takes part. That makes a signal, which binds the large subunit of ribosome and
activates it. In each mRNA the coding sequence of the RNA starts with a start codon, which is
formed by three bases AUG. In prokaryotes the initiation codon is for tRNA, which binds N-
methyl formyl methionine.

The synthesis of the peptide chain starts by the formation of the so called initiation complex.
The formation of the initiation complex is enabled by the helping proteins, which we call the
initiation factors (IF). In prokaryotic cells there are three such factors (IF1, IF2 and IF3), in
eukaryotes there are 11 of such factors and called as eIF.

In prokaryotic cells the IF3 bonds to the 30S ribosomal subunit. Formyl methionine binds as the
first amino acid in the sequence. That is why the binary complex (fMet~tRNA+IF2) is formed
with the help of IF2. By the binding of GTP (guanosine triphosphate) a ternary complex
(fMet~tRNA+IF2+GTP) is formed. After this the small subunit connects with the ternary
complex, to which binds the mRNA forming the preinitiation complex (mRNA+30S+IF3+fMet
~tRNA+IF2+GTP+IF1). Then, after GTP hydrolysis, all three initiation factors release and the
large (50S) ribosomal subunit binds, forming the initiation complex. The initiation complex
consists of both ribosomal subunits, and mRNA bond on the small subunit the. The formation of
the initiation complex in eukaryotic cell is a more complex process.

Four active sites appear in the activated ribosome:

 A – to amino acid enter, bonded to the corresponding tRNA;
 P – peptide, where the amino acids bind by peptide bonds;
 E – for the exit of the newly synthesized protein from the ribosome;
 R – for the exit of the tRNA which gave an amino acid.

Methionine doesn’t go to the A site but directly to the P site, since only fMet~tRNA in
prokaryotes and Met~tRNA in eukaryotes are able to activate the P site – that is why they are
always at the beginning of the synthesized protein.
Activation of the amino acid and its binding to the tRNA

The tRNA has an important role in reading of the code. To the adenine on its 3´-OH acceptor arm
an amino acid is attached.

Amino acids, which are supposed to be build-in into the protein molecule are not able to read the
information in the mRNA – by themselves. This is why the amino acid has to connect to the
tRNA, which transfers it to a ribosome. This connection is carried out by aminoacyl tRNA
synthetase – enzyme forming aminoacyltRNA. Each of the amino acids has its own tRNA,
which is specific for the connection with only one amino acid. Aminoacyl-tRNA synthetase is
specific because it binds to the kind of tRNA, whose anticodon is complementary to the codon.
The lengthening of the peptide chain - elongation

A second complex aminoacyl-tRNA carrying its amino acid enters to the A site. Sequentially
tRNA connects to the mRNA, by the codon-anticodon bond. The aminoacyl tRNA is not able to
bind to the mRNA by itself. A so called elongation factor EF1 and GTP are required. First, a
ternary complex of aminoacyl-tRNA + EF1 + GTP, is formed. If the anticodon in the tRNA is not
complementary to the codon in the mRNA, the binding cannot take place and the aminoacyl-
tRNA is substituted by another one, which is complementary.

The second aminoacyl-tRNA moves from A site to P site of ribosome large subunit and here
located enzyme the peptidyl transferase connects first and second amino acids by a peptide
bond. Then protein elongation factor 2 (EF2) „pushes “the second tRNA. It results to movement
of the mRNA by one triplet – following one towards mRNA 3´ end. The tRNA which brought the
first amino acid is released from ribosome and liberates its space on mRNA for a second tRNA –
which made a space for a third tRNA (carrying third amino acid). Third aminoacyl-tRNA moves
to A site and then to P site, where peptide bond between second amino acid and third one is done.
Then again acts EF2, comes fourth aminoacyl-tRNA etc. The newly synthesized peptide exits
from the ribosome through the E site. The whole process repeats. The ribosome must always
contain two tRNAs; one which is ready to leave and the second one which has just arrived. By
the repetition of these steps the peptide chain is slowly contains all the amino acids, according to
the sequence of codons in the mRNA molecule. This process is called elongation, or the
lengthening of the protein product.

The termination of the synthesis of the polypeptide

The process continues until the mRNA reaches to the stop triplet, which signals the termination
of the polypeptide chain formation. There is no tRNA for the stop triplet, meaning that the last
tRNA remains alone in the ribosome. The outcome is the disintegration of the ribosome and the
termination of translation.

Post translational modifications of the proteins

In eukaryotic cells, initiation and the beginning of translation takes place in the cytoplasm. The
primary protein chain is modified for instance by the removal of the first methionine or various
amino acids, by hydroxylation, glycosylation or phosphorylation, the production of disulfide
bridges within the chain, the formation of tertiary structure of the protein etc. A common
modification is the removal of the part of the peptide, a good example is pro-insulin. The removal
of the polypeptide C, which consists of 33 amino acids, and the connection of the A chain (21
amino acids) and B chain (30 amino acids), functional insulin is formed.