What is Ribosomes?: Its structure and function in translation

Ribosomes:

Ribosomes ( singular= Ribosome) are present either in the cytoplasm or loosely attached to the cell membrane (plasma membrane) of the cell. they are complex structures that contain ribonucleic acid (RNA) which is known as ribosomal RNA (rRNA) and proteins. Proteins of cells form in the ribosomes. Ribosomes of cytoplasm form proteins that remain inside the cell while that is attached to the plasma membrane form proteins that are transported outside the cell.

The cells that have higher rates of protein synthesis contain a larger number of ribosomes. The ribosomes present in the cytoplasm give their granular appearance. Ribosomes cannot be seen by light microscope.

STRUCTURE OF PROKARYOTIC RIBOSOME:

Prokaryotic ribosomes are different from eukaryotic ribosomes. These ribosomes are smaller than the eukaryotic ribosomes and are known as 70S ribosomes. Their molecular weight is approximately 2.7 million and have dimensions of approximately 15nm by 20 nm. The process of protein synthesis from mRNA explains the structure of ribosomes..  

Prokaryotic ribosomes are made up of two subunits. The larger subunit is the 50S while the smaller subunit is 30S (here S has the same values that stand for the Svedberg unit). This unit shows the sedimentation rate at ultra-high-speed centrifugation. The sedimentation rate depends on the weight, size, and shape of the particle.

The mRNA- binding site binds a sequence near the 5 prime ends of m RNA for translation of the first codon. The binding sites are located at or near large and small subunits.

The smaller subunit has a molecular weight of 900,000 and has 21 ribosomal proteins and one 16S ribosomal RNA. The ribosomal RNA molecules are capable of folding into many different secondary structures.

16S and 23S rRNAs have 30 post transcriptional modified nucleotides. All of these nucleotides form a group at the A site surrounding the anticodon loop of the P site. These modified nucleotides (bases) in 23S rRNA gather around peptidyl transferase at the center of the 50S subunit and these modifications are the reasons for fine interactions between rRNA and tRNA.

The larger subunit has a molecular weight of 1.6 million and has 31 ribosomal proteins and ribosomal RNA species (one copy of both 23S and 5S rRNA).  So a ribosome consists of 52 ribosomal or r-proteins and three different types of RNA.

STRUCTURE OF EUKARYOTIC RIBOSOME:

The eukaryotic ribosome is larger than the prokaryotic ribosome (the 80S). It also has two subunits; the larger one is the 60S and the smaller one is the 40S. The smaller subunit has 18S ribosomal RNA and the larger subunit has 23S ribosomal RNA.

FUNCTION OF RIBOSOMES:

Ribosomes are involved in protein synthesis. In eukaryotes, ribosomes get their order from the nucleus to form proteins. In the nucleus, transcription occurs (mRNA is formed) and this mRNA goes to the ribosome where the process of translation occurs (proteins are formed).

In prokaryotic cells, the nucleus is not present mRNA is formed in the cytoplasm and also translated into the cytoplasm by ribosomes.

BINDING SITES OF RIBOSOMES:

The ribosome has three tRNA binding sites:

1. A site: aminoacyl tRNA binding site or also called decoding site
2. P sit: peptidyl binding site
3. E sit: exit site

TRANSLATION IN PROKARYOTES:

It occurs in three steps initiation, elongation, and termination.

Initiation factors:

It has three initiation factors: IF-1, IF-2, and IF-3.

• IF-1: it helps in the dissociation of a large and small unit of the ribosome.
• IF-2: it is the t-RNA with N-formyl-methionine attached with IF-2.
• IF-3: it binds to the small subunit of a ribosome and prevents the pre-mature binding of both subunits of Ribosomes.

In prokaryotes, protein synthesis is initiated by a modified methionine residue i.e. N-formyl methionine-tRNA. Chain initiation begins with the formation of  two complexes:

1. 1. IF-2 & N-formyl methionine t-RNA
2. 2. m-RNA, 30S subunit, IF-3

Prokaryotic mRNAs contain a Shine Dalgarno Sequence. Shine Dalgarno sequence is complementary to nucleotide sequence at 3 prime ends of ribosomal RNA of the 30 S subunit.

Interaction between these complementary sequences enhances the attachment of the 30S subunit to the AUG initiator codon. Both complexes combine with each other with IF-1 and 1 molecule of GTP.

Then 50 S subunit gets added to the complex structure and IF-3 is released. The addition of the 50S subunit utilizes the GTP which in turn activates the release of IF-1 and IF-2 factors.

The addition of the 50S subunit to the complex places the N-formyl met tRNA in the P site directly with the anticodon of the tRNA aligned with the AUG codon of mRNA. When the AUG initiator codon is positioned in the P site, the second codon in the m-RNA positions in such a way that it corresponds to the incoming aminoacyl-t RNA in the A site.

ELONGATION FACTORS:

The most common elongation factors are EF-Tu, EF-Ts, and EF-G. Their function is to facilitate translation from the formation of the first peptide bond to the last peptide bond of the growing polypeptide chain.

Elongation begins with the second aminoacyl t-RNA at the ribosomal aminoacyl (A) site. The t-RNA travels to the A site by the elongation factor EF-Tu which also carries bound GTPs. As the t-RNA binds, the GTPs are hydrolyzed and EF-Tu is released.

A peptide bond formed between the N-f-met-t RNA at the P site and 2^nd aminoacyl t-RNA at the A site; catalyzed by the peptidyl transferase. This is the transfer of N-f-met.to aminoacyl-tRNA at the A site forming a peptidyl tRNA at that position and leaving an uncharged tRNA at the P site. Peptidyl tRNA is translocated to the P site and uncharged tRNA is translocated to the E domain

A site is now unoccupied; new aminoacyl-Trna bind to A site and this process continues until the last peptide bond forms.

TERMINATION FACTORS:

1. RF-1: it recognizes UAG and UAA codons
2. RF-2:  it recognizes UGA and UAA codons
3. RF-3: it facilitates the removal of RF-1 from ribosomes following peptide release.

When a stop codon (UAG, UAA, UGA) arrives at the A site, it is recognized and bound by a protein release factor. RFs bind to the termination codon at A site and stimulate hydrolysis of the bond between tRNA and polypeptide chain at P site, resulting in the release of the complete polypeptide from ribosomes.

Termination is completed by the release of mRNA molecules from ribosomes and the dissociation of ribosomes into their subunits.

TRANSLATION IN EUKARYOTES

Initiation Factors:

• eIF-2:    Binds tRNA^Met to 40S subunit
• eIF-1A: First factor that binds to 40S subunit
• eIF-3:    Binds to 40S subunit, prevents 60S subunit from binding to it
• eIF-4A:  Remove secondary structures in mRNA to promote 40s binding to mRNA
• eIF-4B: Helps in to find start codon AUG
• eIF-4E: Help in the recognition of the 5” cap
• eIF-4G: Helps in the recognition of 3’ tail
• eIF-5:    Stimulates 60S subunit binding to the 40S pre-initiation complex
• eIF-6:    Binds to 60S subunit, helps prevent 40S subunit from binding to it

The initiation is divided into the following steps :

1. Activation of mRNA
2. Ternary Complex
3. Formation of 43S preinitiation complex.
4. Formation of 48S initiation complex.
5. Formation of 80S initiation complex.

Elongation:

Ribosomes elongate the polypeptide chain by the sequential addition of amino acids.  Elongation is a cyclic process involving certain elongation factors (EFs).
 It is divided into three steps.

1.  Binding of Aminoacyl t-RNA to A-site.
2. Peptide bond formation.
3.  Translocation

Elongation Factor:

• eEF1A:  it binds to charged aminoacyl tRNA and brings it to the A site
• eEF1B, eEFG, eFFG : they will help in the replacement and substitution of GDP to GTP
•  eEF2: it helps in Translocation

Termination:

It has two stages:

• Cleavage of a growing chain
• Recycling factor

One of the stop or termination signals (UAA, UAG, and UGA) terminates the growing polypeptide.
• When the ribosome encounters a stop codon, there is no tRNA available to bind to the A site
– instead, a release factor binds to it.
• Ribosome recycling occurs in eukaryotes.

References:

https://www.britannica.com/science/ribosome

https://www.nature.com/scitable/topicpage/ribosomes-transcription-and-translation-14120660/