Introduction:
Protein synthesis is one of the most vital biological processes, where cells produce new proteins based on the genetic information carried by messenger RNA (mRNA). This process is known as translation.
Translation occurs in the cytosol, specifically on ribosomes, and is guided by the sequence of codons present on mRNA. The overall process is similar in both prokaryotes and eukaryotes, although eukaryotic translation involves more factors and additional steps of regulation and control.
Basic Requirements for Translation:
Several key components are necessary for successful protein synthesis:
- mRNA: Carries the genetic message from DNA.
- tRNA: Transfers specific amino acids to the ribosome.
- Ribosomes: Act as the site of protein synthesis.
- Energy molecules: ATP and GTP provide energy for various steps.
- Protein factors: Such as initiation factors, elongation factors, and termination factors.
Stages of Eukaryotic Translation:
The process of protein synthesis in eukaryotes takes place in four main stages:
- Activation of amino acids
- Initiation
- Elongation
- Termination
1. Activation of Amino Acids:
This step takes place in the cytosol. Each of the 20 amino acids is attached to its specific transfer RNA (tRNA) molecule. This process is catalyzed by an enzyme called aminoacyl-tRNA synthetase and requires energy from ATP.
The result is the formation of aminoacyl-tRNA, a molecule that can recognize a codon on the mRNA through its anticodon region and bring the correct amino acid for protein assembly.
2. Initiation:
Initiation is the most complex part of protein synthesis and can be divided into four key steps:
- Ribosomal dissociation
- Formation of 43S pre-initiation complex
- Formation of 48S initiation complex
- Formation of 80S initiation complex
Step 1: Ribosomal Dissociation:
Eukaryotic ribosomes are 80S particles made up of two subunits — 40S and 60S. The 40S subunit binds to initiation factors (eIF3 and eIF-1A), which help separate the ribosome into its two subunits for initiation to begin. The ribosome contains two tRNA binding sites: A site (aminoacyl) and P site (peptidyl).
Step 2: Formation of 43S Pre-Initiation Complex:
Initiation begins with GTP binding to initiation factor eIF2. This GTP–eIF2 complex then binds to Met-tRNAiMet (initiator tRNA), forming a ternary complex. In prokaryotes, the first amino acid is N-formylmethionine (fMet), but in eukaryotes, it is simply methionine.
This ternary complex then binds to the 40S ribosomal subunit, forming the 43S pre-initiation complex, stabilized by eIF3 and eIF-1A.
Step 3: Formation of 48S Initiation Complex:
The 43S complex now binds to mRNA to form the 48S initiation complex. The 5′ end of most eukaryotic mRNAs carries a methylguanosine cap that helps mRNA attach to the ribosome via the cap-binding protein complex (eIF4) and requires ATP. In prokaryotes, mRNA binds to the ribosome using the Shine–Dalgarno sequence.
Step 4: Formation of 80S Initiation Complex:
The large 60S ribosomal subunit then joins the 48S initiation complex to form the 80S initiation complex. This step requires hydrolysis of GTP (bound to eIF2) by eIF5, which also releases the initiation factors for recycling.
At this point, the initiator tRNA occupies the P site and pairs with the start codon (AUG) on mRNA, ready to begin elongation.
3. Elongation:
Elongation is the process of adding amino acids one by one to the growing polypeptide chain. It consists of three steps:
Step 1: Binding of Aminoacyl-tRNA to the A Site:
The next aminoacyl-tRNA, corresponding to the next codon on mRNA, binds to the A site of the ribosome. This step requires elongation factor eEF-1α and GTP, which are then released as eEF-1α–GDP and phosphate.
Step 2: Formation of Peptide Bond:
A new peptide bond forms between the amino acid at the A site and the growing peptide chain attached to the tRNA at the P site. This reaction is catalyzed by peptidyl transferase, a ribozyme activity of the ribosome. As a result, the peptide chain shifts to the tRNA in the A site.
Step 3: Translocation:
The ribosome moves along the mRNA by one codon toward the 3′ end, a process called translocation. The dipeptidyl-tRNA moves from the A site to the P site, and the empty tRNA exits the P site. This process is driven by elongation factor eEF2 and the hydrolysis of GTP.
The ribosome is now ready for the next aminoacyl-tRNA, and the elongation cycle continues until a stop codon appears.
4. Termination:
Elongation continues until the ribosome encounters one of the three termination (stop) codons — UAA, UAG, or UGA — on the mRNA.
- These codons do not code for any amino acid. Instead, they are recognized by release factors that signal the end of translation.
- In eukaryotes, there is a single release factor called eRF, while prokaryotes have three (RF-1, RF-2, and RF-3).
- The release factor, together with GTP and peptidyl transferase, hydrolyzes the bond between the growing polypeptide and the tRNA in the P site, releasing the newly synthesized protein.
After the polypeptide chain is released, the mRNA detaches from the ribosome, and the ribosome dissociates into its 40S and 60S subunits for reuse.
Detailed Notes:
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