The translation in eukaryotes is the biological process by which protein is synthesized from the information coded on the molecule of messenger RNA (mRNA). It can also be defined as the process in which nucleotides sequence on the messenger RNA molecule helps to incorporate amino acids into protein.
COMPONENTS REQUIRED FOR PROTEIN SYNTHESIS IN EUKARYOTIC TRANSLATION
Four primary components are required to form protein from the nucleotide sequence or information coded on messenger RNA.
- mRNA: it contains DNA sequence or genetic information in form of codons to form proteins
- tRNA: it behaves like an adapter molecule between amino acids or codons, required to carry amino acids to ribosomes to synthesize protein
- 80S ribosomes (60S and 40S): these are protein synthesis machinery which means proteins are formed in ribosomes.
- Enzymes: these are required for the correct attachment of amino acids to tRNA molecule. Two enzymes are involved in this process:
- aminoacyl tRNA synthetase: catalyze the attachment of tRNA molecule to its respective amino acid
- peptidyl transferase: catalyzes the sequential transfer of amino acid to the growing chain.
SITE OF TRANSLATION
In prokaryotes, transcription and translation occur simultaneously and both mechanisms occur in the cytoplasm. While in eukaryotes, transcription occurs in the nucleus and translation occurs in the cytoplasm.
RIBOSOMES ROLE IN TRANSLATION
The smaller subunit of ribosome (the 40S) binds with the mRNA and the larger subunit (60S) provides the enzymatic activity. The structure of the ribosome also provides three sites for the binding of tRNA molecules.
- A site
- P site
- E site
ACTIVATION OF TRANSFER RNA
Activation of transfer RNA is very important in the translational process. It involves two steps:
- Activation of amino acid
- Attachment of Amino acid to tRNA
MECHANISM OF TRANSLATION
Just like the prokaryotic process of translation, translation in eukaryotes also occurs in three steps and each step is aided by its respective factors.
The process of translation initiation in eukaryotes is complex and it involves four steps and ten eukaryotic initiation factors.
STEPS OF INITIATION:
- Dissociation of ribosome
- Formation of 43S preinitiation complex
- Formation of 48S initiation complex
- Formation of 80S initiation complex
- eIF-2: it binds the tRNAMet with 40S subunit
- eIF-1A: it is the first factor that binds with the 40S subunit
- eIF-3: it binds to the 40S subunit thus prevents 60S subunit from binding to it
- eIF-4A: it removes the secondary structures in mRNA for the 40S binding to mRNA
- eIF-4B: it helps in finding of start codon AUG
- eIF-4E: it helps in the recognition of a 5’ cap
- eIF-4G: it helps in the recognition of 3’ tail
- eIF-4F: it binds to cap of messenger RNA
- eIF-5: it stimulates the 60S subunit binding with the 48S pre-initiation complex
- eIF-6: it binds to 60S subunit and helps to prevent 40S subunit from binding to it
DISSOCIATION OF RIBOSOME
The 80S ribosome of eukaryotic cell dissociates into 60S and 40S subunits. Then eukaryotic initiation factors eIF-3 and eIF-1A binds to the 40S ribosomal subunit to prevent the 60S subunit reassociation with 40S subunit.
FORMATION OF 43S PREINITIATION COMPLEX
In this step, a ternary complex formed that contained met-Trna and eukaryotic IF-2 bound to the GTP molecule. This is then attaches to the 40S subunit and form 43S pre-initiation complex. The eukaryotic IF-3 and IF-1A help to stabilize this complex.
FORMATION OF 48S INITIATION COMPLEX
Formed 43S pre-initiation complex binds to messenger RNA and form 48S initiation complex. Also, eukaryotic IF-4F complex formed by the association of eukaryotic IF-4A, IF-4G with the eukaryotic IF-4E. The eukaryotic IF-4F is also known as the cap binding protein because it binds to the cap of messenger RNA. After that complex messenger RNA structure will reduce by the binding of eukaryotic IF-4A and IF-4B to messenger RNA.
This messenger RNA will then be transferred to the 43S complex. Energy in the form of ATP is required for the proper association of 43S pre-initiation complex with messenger RNA. Messenger RNA will be scanned by the ribosomal initiation complex to identify the accurate initial codon. The initiation codon is 5’-AUG.
FORMATION OF 80S INITIATION COMPLEX
The 80S initiation complex will be formed by the binding of the 48S initiation complex with 60S ribosomal subunit. This binding requires the hydrolysis of GTP molecule and it is aided by the eukaryotic IF-5. After the formation of 80S initiation complex, the initiation factors bound to 48S initiation complex will be released and recycled.
Ribosomes will elongate the polypeptide chain by the addition of amino acids in a sequence. The sequence of amino acids will be determined by the codons’ order on the specific messenger RNA. Elongation is a cyclic process that involves many elongation factors (EFs).
The elongation process is divided into three major steps:
- Binding of Aminoacyl t-RNA to A-site.
- Peptide bond formation.
- eEF1A: it binds to the charged aminoacyl tRNA and brings it to A site
- eEF1B, eEFG, and eFFG: these factors help in the replacement and substitution of GDP to GTP
- eEF2: it helps in the process of translocation
BINDING OF AMINOACYL T-RNA TO A-SITE
The P-site of the 80S initiation complex has met-Trna and its A site is free. The aminoacyl-Trna will be placed at the A site. Eukaryotic EF-1A is involved in this process and a supply of energy is required in the form of GTP. Accurate codons are recognized on messenger RNA. Then GDP and EF-1A are recycled and another aminoacyl-tRNA is placed at the A site.
PEPTIDE BOND FORMATION
The peptide bond formation is catalyzed by the enzyme peptidyl transferase. This enzyme performs its activity on 28S RNA of the 60S subunit of the ribosome. Hence peptide bond formation occurs by the attachment of the growing peptide chain with the transfer RNA at the A-site.
The ribosome will then move towards the 3’ end to the next codon of messenger RNA. This process is known as translocation. as it involves the movement of a growing peptide chain from A-site to P-site. Eukaryotic EF-2 and GTP are required in the process of translocation. GTP will be hydrolyzed and provides energy to messenger RNA. EF-2 and GTP can also be recycled for translocation.
There are two stages of termination in eukaryotic cells:
- Cleavage of a growing chain
CLEAVAGE OF GROWING CHAIN
One of the stop codon or termination signals (UAA, UAG, and UGA) will terminate the growing polypeptide. In eukaryotes, all of three stop codons are recognized by one eukaryotic release factor eukaryotic RF-1. Eukaryotic RF-3 helps to stimulate termination. When the ribosome comes in contact with a stop codon, there will be no tRNA available for binding to the A site and instead of it a release factor will bind to it. After the binding of the release factor, the ribosome will fall apart into the following components:
- Small and large ribosomal subunits will be released
- Peptidyl product will be freed
- Transfer RNA will also be released
Recycling of ribosomes occurs in eukaryotes. When the polypeptide and release factors are released, the ribosomes are still bound to the messenger RNA and have the deacylated transfer RNA in the P and E sites. For the new polypeptide synthesis, these mRNA and tRNA must be separated and ribosomes must also be dissociated. All of these processes are collectively known as ribosome recycling.
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