Protein Synthesis

Protein synthesis
1. DNA unwinds
2. mRNA copy is made of one of the DNA strands.
3. mRNA copy moves out of nucleus into cytoplasm.
4. tRNA molecules are activated as their complementary amino acids
are attached to them.
5. mRNA copy attaches to the small subunit of the ribosomes in
cytoplasm. 6 of the bases in the mRNA are exposed in the
ribosome.
6. A tRNA bonds complementarily with the mRNA via its anticodon.
7. A second tRNA bonds with the next three bases of the mRNA, the
amino acid joins onto the amino acid of the first tRNA via a
peptide bond.
8. The ribosome moves along. The first tRNA leaves the ribosome.
9. A third tRNA brings a third amino acid
10. Eventually a stop codon is reached on the mRNA. The newly
synthesised polypeptide leaves the ribosome.
Overview
Transcription 1
(making a mRNA copy of DNA)

•The part of the DNA molecule (the gene) that the cell wants the
information from to make a protein unwinds to expose the bases.
•Free mRNA nucleotides in the nucleus base pair with one strand
of the unwound DNA molecule.
 Transcription 2

•The mRNA copy is made with the help of RNA polymerase. This enzyme
joins up the mRNA nucleotides to make a mRNA strand.
•This mRNA strand is a complementary copy of the DNA (gene)
•The mRNA molecule leaves the nucleus via a nuclear pore into the
cytoplasm
tRNA –
pick up their specific amino acids
from the cytoplasm
Transfer RNA Molecules Have a Common Design

1. Each is a single chain containing between 73 and 93

ribonucleotides (~25 kd).

2. They contain many unusual bases, typically between 7 and 15 per

molecule. Some are methylated or dimethylated
3. About half the nucleotides in tRNAs are base-paired to form double
helices.

Five groups of bases are not base paired ,in this way:

the 3 CCA terminal region, which is part of a region called the acceptor stem;

the TyC loop, which acquired its name from the sequence ribothymine-
pseudouracil-cytosine;

the "extra arm," which contains a variable number of residues;

the DHU loop, which contains several dihydrouracil residues;

and the anticodon loop.

The structural diversity generated by this combination of helices and

loops containing modified bases ensures that the tRNAs can be uniquely
distinguished, though structurally similar overall.
4. The 5 end of a tRNA is phosphorylated. The 5 terminal residue is
usually pG.

5. The activated amino acid is attached to a hydroxyl group of the

adenosine residue located at the end of the 3 CCA component of the
acceptor stem. This region is single stranded at the 3 end of mature
rRNAs.

6. The anticodon is present in a loop near the center of the sequence.

General Structure of tRNA Molecules
Amino Acids Are First Activated by Adenylation

The activation reaction is catalyzed by specific aminoacyl-tRNA synthetases,

which are also called activating enzymes.
The first step is the formation of an aminoacyl adenylate from an amino acid
and ATP.
The next step is the transfer of the aminoacyl group of aminoacyl-AMP to a
particular tRNA molecule to form aminoacyl-tRNA.

The sum of these activation and transfer steps is

Translation - animation
mRNA
attaches to
small
ribosomal
subunit
 A Ribosome Is a Ribonucleoprotein Particle (70S) Made
of a Small (30S) and a Large (50S) Subunit

An E. coli ribosome is a ribonucleoprotein assembly with a mass of about 2700 kd, a

diameter of approximately 200 Å, and a sedimentation coefficient of 70S. The
20,000 ribosomes in a bacterial cell constitute nearly a fourth of its mass.

A ribosome can be dissociated into a large subunit (50S) and a small subunit (30S) .

The 30S subunit contains 21 different proteins (referred to as S1

through S21) and a 16S RNA molecule.

The 50S subunit contains 34 different proteins (L1 through L34) and two RNA
molecules, a 23S and a 5S species.

A ribosome contains one copy of each RNA molecule, two copies of the L7 and L12
proteins, and one copy of each of the other proteins. The L7 protein is identical
with L12 except that its amino terminus is acetylated. Only one protein is common
to both subunits: S20 is identical with L26.
Translation - outline
 Messenger RNA Is Translated in the 5 -to-3
Direction

An important feature of prokaryotic gene expression is that translation

and transcription are closely coupled in space and time.

Many ribosomes can be translating an mRNA molecule simultaneously.

This parallel synthesis markedly increases the efficiency of mRNA

translation.

The group of ribosomes bound to an mRNA molecule is called a

polyribosome or a polysome
Bacterial Protein Synthesis Is Initiated by Formylmethionyl
Transfer RNA

Protein synthesis in bacteria starts with N-formylmethionine (fMet).

A special tRNA brings formylmethionine to the ribosome to initiate protein
synthesis. This initiator tRNA (abbreviated as tRNAf) differs from the one that
inserts methionine in internal positions (abbreviated as tRNAm).
In approximately one-half of E. coli proteins, Nformylmethionine is removed when
the nascent chain is 10 amino acids long.
Methionine is linked to these two kinds of tRNAs by the same amino-acyl-tRNA
synthetase.

A specific enzyme then formylates the amino group of methionine that is

attached to tRNAf.

The activated formyl donor in this reaction is N 10-formyltetrahydrofolate .

It is significant that free methionine and methionyl-tRNAm are not substrates

for this transformylase.
Formylation of Methionyl-tRNA
Translation.
mRNA used to make polypeptide chain
(protein)
1.

•First the mRNA attaches itself to a ribosome (to the small subunit).
•Six bases of the mRNA are exposed.
•A complementary tRNA molecule with its attached amino acid (methionine) base
pairs via its anticodon UAC with the AUG on the mRNA in the first position P.
•Another tRNA base pairs with the other three mRNA bases in the ribosome at
position A.
•The enzyme peptidyl transferase forms a peptide bond between the two amino
acids.
 Translation 2

The ribosome moves along the mRNA to the next codon (three bases).
The second tRNA molecule moves into position P.
Another tRNA molecule pairs with the mRNA in position A bringing its amino
acid.
A growing polypeptide is formed in this way until a stop codon is reached.
 End of Translation

A stop codon on the mRNA is reached and this signals the ribosome to leave
the mRNA. A newly synthesised protein is now complete!
Translation
mRNA to Polypeptide