Faculty of Medicine – Department of Medicine – Course of Cytology –

First year of medicine – Academic year 2025/2026

RIBOSOMES
Dr. R. KHAROUBI

I. GENERAL INTRODUCTION
Ribosomes are complex and fascinating cellular structures responsible for protein synthesis
in all living organisms. They represent a critical step in the transfer of genetic information,
from DNA to protein via RNA. In essence, they decipher the information encoded in DNA by
using messenger RNA (mRNA) to assemble a sequence of amino acids corresponding to the
protein to be produced. This process, known as "translation," is fundamental to proper
cellular function, which explains the long-standing research interest in ribosomes.

Ribosomes are also of considerable interest in biomedicine : on one hand, they are a primary
target for several clinically important antibiotics; on the other hand, the regulation of protein
synthesis plays a crucial role in major biological processes such as cell growth,
differentiation, as well as in viral infections and various diseases, including cancers.

II. HISTORICAL BACKGROUND

The concept of ribosomes dates back to the 1950s and 1960s, thanks to the first observations
made using electron microscopy. Before this, protein synthesis was poorly understood.

In 1955, the molecular biologist George Palade, through his electron microscopy research, was
one of the first to identify the structures involved in this synthesis. He initially called them
"granules" before they were renamed "ribosomes."

During the 1960s, research confirmed that ribosomes are present in all living cells, from
prokaryotes to eukaryotes, and that they are essential for the translation of genetic information.

Our understanding of ribosomes has advanced considerably thanks to technological

breakthroughs such as X-ray crystallography, which made it possible to determine their
structure in detail. This major advance was made possible by the work of researchers like
Venkatraman Ramakrishnan, Thomas A. Steitz, and Ada Yonath, who were awarded the Nobel
Prize in Chemistry in 2009 for their discoveries regarding the structure and function of the
ribosome.

1
III. STRUCTURE AND COMPOSITION

Ribosomes are composed of two distinct subunits, a large and a small one, which assemble to
carry out the translation of genetic information. Their size and composition vary depending on
whether they are found in prokaryotic or eukaryotic cells.

1. Small Subunit (30S subunit in prokaryotes, 40S in eukaryotes) :

- This subunit is responsible for recognizing and initiating translation by binding to messenger
RNA (mRNA).
- It is smaller and contains a significant amount of ribosomal RNA (rRNA) as well as several
proteins.
2. Large Subunit (50S subunit in prokaryotes, 60S in eukaryotes) :
- It plays a role in the elongation of the polypeptide chain by catalyzing the formation of peptide
bonds between amino acids.
- This subunit is larger and also contains a substantial quantity of rRNA and proteins.

Ribosomes are composed of ribonucleoproteins : ribosomal proteins and ribosomal RNA

(rRNA). The number of proteins and the proportion of rRNA vary between prokaryotes and
eukaryotes :

• In Prokaryotes (e.g., E. coli):

- The ribosome is composed of a small subunit (30S) and a large subunit (50S), totaling
approximately 70S.
- The small subunit (30S) contains the 16S rRNA and approximately 21 proteins.
- The large subunit (50S) contains the 23S and 5S rRNA and approximately 34 proteins.

• In Eukaryotes (e.g., human cells) :

- The ribosome is larger, composed of a small subunit (40S) and a large subunit (60S), totaling
approximately 80S.
- The small subunit (40S) contains the 18S rRNA and approximately 33 proteins.
- The large subunit (60S) contains the 28S, 5.8S, and 5S rRNA and approximately 49 proteins.
- Ribosomal proteins play a crucial role in the proper functioning of ribosomes by facilitating
the translation of the genetic information present in mRNA. They are mainly located on the
cytosolic surface of the ribosome, rather than at the interface between the two subunits.

It is important to note that Svedberg units are not additive; the sum of the two subunits does not
equal the value of the complete ribosome (e.g., 30S + 50S ≠ 80S, but 70S).

IV. ORGANISATION AND BINDING SITE

The ribosome contains an mRNA binding site on the small subunit and three tRNA binding sites
located on the large subunit.

1. mRNA Binding Site (within the small subunit) : Located on the small subunit of the
2
ribosome. It allows the mRNA to attach to the ribosome and determines the reading of the
genetic information contained within the mRNA.
2. The A Site (Aminoacyl site) : The entry site for aminoacyl-tRNAs (tRNAs charged with an
amino acid). This is where the tRNA corresponding to the codon on the mRNA binds during
translation.
3. The P Site (Peptidyl site) : The site where the tRNA carrying the growing polypeptide chain
binds. The formation of the peptide bond between the amino acid in the A site and the
polypeptide chain in the P site occurs here.
4. The E Site (Exit site) : This is the site through which deacylated tRNAs (having transferred
their amino acid to the growing polypeptide) exit the ribosome.

Within the ribosome, several protein factors bind to enable protein synthesis. These factors are
essential for each stage of the translation process :

- First, Initiation Factors (eIFs) : These bind to the ribosome to help assemble the initiation
complex. They play a crucial role in the correct initiation of translation.
- Next, Elongation Factors (eEFs) : These intervene to facilitate the entry of transfer RNAs
(tRNAs) into the A site of the ribosome and the movement (translocation) of the ribosome
along the mRNA.
- Finally, Release Factors (eRFs) : These bind to the ribosome when it encounters a stop
codon. This allows the release [of the complete polypeptide chain from the ribosome].

V. RIBOSOME BIOGENESIS

- The synthesis and processing of ribosomal components occur not only in the nucleolus but
also in the nucleoplasm and cytoplasm of eukaryotic cells.

- The first step in biogenesis is the transcription of the genes coding for rRNAs. This takes
place in the nucleolus. The genes that code for the 18S, 5.8S, and 28S rRNAs in eukaryotes
are transcribed by RNA polymerase I, while the 5S rRNA is transcribed by RNA polymerase
III.

- Once the rRNA is transcribed, it undergoes several modifications, including methylation and
pseudouridylation, which are important for its stability and functions. These modifications
are carried out by enzymatic complexes within the nucleolus. Subsequently, the rRNAs
undergo cleavage steps to generate the mature forms of rRNA.

- Ribosomal proteins are encoded by genes in the nucleus and synthesized in the cytoplasm.
Once synthesized, these proteins are transported back into the nucleus, and then into the
nucleolus, where they assemble with the rRNAs to form the two ribosomal subunits.

- The ribosomal subunits are assembled in the nucleolus and undergo final adjustments before
being transported into the cytoplasm. The small and large subunits are exported individually
through the nuclear pores into the cytoplasm.

3
VI. FUNCTIONS OF RIBOSOMES : PROTEIN SYNTHSIS

- As with mRNA synthesis, protein synthesis can be divided into three phases: initiation,
elongation, and termination. The translation process is similar in prokaryotes and eukaryotes.
- The key players in translation are ribosomes, tRNAs that carry amino acids, and mRNA
(messenger RNA).
- In eukaryotes, ribosomes are either free in the cytoplasm or bound to the endoplasmic
reticulum (referred to as the rough endoplasmic reticulum), and translation occurs in the
cytoplasm.
- In prokaryotes, transcription and translation are coupled (simultaneous), whereas in
eukaryotes, translation occurs in the cytoplasm using mature mRNA.
- When several ribosomes are translating the same mRNA molecule, this structure is called a
polysome.
- The different stages of protein biosynthesis require energy provided by the hydrolysis of
GTP or ATP.

1-Initiation

1/ Formation of the Initiation Complex

• Initiation of protein synthesis requires the presence of :
✓ The small subunit (SSU) 40S
✓ The initiator tRNA, which brings the first amino acid in the form of methionine linked to the
tRNA (Met-tRNAi^Met).
✓ The mRNA.
✓ Initiation factors.
• The initiation phase ends with the association of the large subunit (LSU) with the small subunit
(SSU), thus forming a complete initiation complex.
• In eukaryotes, the 5' end carries a cap (Cap Binding Protein, CBP) consisting of
methylguanosine molecules. The SSU, assisted by the cap-binding complex eIF4F (composed of
eIF4E, eIF4G, and eIF4A), recognizes the cap and then moves along the mRNA molecule until it
reaches the sequence 5'-CCACCAUGC-3', which contains AUG. The translation of the message
carried by the mRNA begins at the AUG initiation codon.

- First Stage of Initiation

• eIF2 binds to GTP.
• An initiator tRNA carrying a methionine molecule (Met-tRNAi^Met) is recognized by eIF2.

- Second Stage of Initiation

• This stage requires the presence of the 40S SSU, which is associated with eIF3 as well as eIF1
and eIF1A. This complex of anti-association factors is located on the face of the SSU that
contacts the LSU, thereby physically blocking the premature binding of the LSU.
• A complex consisting of eIF2-Met-tRNAi^Met-GTP is formed. This complex, associated with
the 40S SSU and the factors eIF3, eIF1, and eIF1A, constitutes the 43S pre-initiation complex.

4
Next, this complex binds to the mRNA to form the 48S pre-initiation complex. The formation of
this complex depends on the presence of the cap, eIF4E, eIF4G, eIF4A, and eIF4B, which enable
the mRNA to bind to the SSU.
• The SSU moves along the mRNA molecule (scanning), over approximately one hundred
nucleotides, in order to find the first AUG start codon.

- Binding of the Large Subunit

• Another initiation factor, eIF5, interacts with the pre-initiation complex and triggers the
hydrolysis of GTP bound to eIF2 ; eIF2-GDP and eIF3, as well as eIF1 and eIF1A, are released
from the SSU.
• Factor eIF5B (a GTPase) then facilitates the association of the LSU with the complex, thus
forming the complete ribosome (80S) which contains the mRNA and the initiator tRNA correctly
positioned in the P site.
• The initiation complex thus formed is ready to function.

2- Elongation

• The elongation phase is characterized by the lengthening of the synthesized protein molecule
and involves elongation factors (EF). It consists of three successive steps :
✓ Binding of an aminoacyl-tRNA.
✓ Formation of a peptide bond.
✓ Ribosomal translocation.

a- Binding of the aminoacyl-tRNA-EF1A-GTP Complex

• A second aminoacyl-tRNA-EF1A-GTP complex enters the A site, regardless of the amino acid
it carries. For it to bind to the mRNA, this aminoacyl-tRNA must be associated with an
elongation factor EF1A bound to GTP, and the anticodon of the tRNA in this complex must be
complementary to the mRNA codon located in the A site.
• If the anticodon and codon are complementary, GTP is hydrolyzed, and the complex releases
EF1A-GDP, which is immediately rephosphorylated into EF1A-GTP.

b- Formation of a Peptide Bond

• The Met-tRNAi is bound to the P site. The next amino acid, whose nature is specified by the
following codon located in the A site, binds to this site.
• The selection of the aminoacyl-tRNA depends on elongation factors: EF1A, which forms a
complex with GTP and the aminoacyl-tRNA. A new complex positions itself in the A site (aa-
tRNA-EF1A-GTP).
• A peptidyl transferase hydrolyzes the Met-tRNAi. The result of this hydrolysis is the transfer of
methionine onto the amino acid (aa1) of the aminoacyl-tRNA located in the A site. This reaction
does not require external energy provided by ATP or GTP: the energy of the methionyl-tRNA
ester bond is sufficient to form a peptide bond between methionine and aa1.
• The tRNA molecule in the A site remains attached at one end to its complementary codon on
the mRNA and at the other end to the methionine-aa1 dipeptide.

5
c- Translocation
• The mRNA and the peptidyl-tRNA (tRNA-methionine-aa1) must be moved so that the mRNA
can place the next codon into the P site and the A site becomes free to receive another
aminoacyl-tRNA.
• The ribosome undergoes a translocation of three nucleotides toward the 3' end of the mRNA.
This step requires GTP hydrolysis catalyzed by elongation factor EF2.
• The newly formed peptidyl-tRNA thus moves from the A site to the P site.
• The arrival of a new aminoacyl-tRNA at the A site leads to the expulsion of the [deacylated]
tRNA, thereby allowing a new cycle to begin.

d- Succession of Elongation Cycles

• The A site is ready to receive an aminoacyl-tRNA-EF1A-GTP complex, provided that the
anticodon of the aminoacyl-tRNA is complementary to the third codon. After the third tRNA
binds to the A site, the dipeptide from the tRNA bound to the P site moves and combines with
the third amino acid. The tRNA from the A site is ejected.
• The ribosome moves by one codon to bring the tripeptide into the P site.
• The now vacant A site is available to receive a fourth aminoacyl-tRNA.
• The succession of similar cycles will result in the synthesis of the protein.

3-Termination

• Translation is completed when a stop codon (UAA, UAG, UGA) appears in the A site
recognition site : no tRNA possesses an anticodon complementary to these three codons.
• A binding factor called RF (Release Factor) binds to the ribosome in the form of an RF-GTP
complex. The peptidyl-tRNA ester bond is then cleaved by peptidyl transferase, which acts as a
hydrolase.
• The peptide is released from the tRNA: this reaction utilizes the energy provided by GTP
hydrolysis. The two subunits separate, and the tRNA and RF-GDP are released into the cytosol.

6
 Figure 01. Structure of ribosomes

Figure 02. Eucaryotic Ribosome VS Procaryotic Ribosome

7
Figure 03. Ribosome biogenesis

8
Figure 04. Translation step

9
 Figure 05. Elongation step

Figure 06. Protein biosynthesis

10