Mechanism of Protein Synthesis

Protein synthesis involves two steps:

1. The activation and charging reaction.

2. The transfer reaction.

1. The activation and charging reaction:

Amino acids in the cytoplasm occur in an inactive stage and they cannot take part in protein synthesis. Hence, these are activated by giving them energy. The activation is facilitated through ATP. In this reaction the carboxyl group of the amino acids react with ATP forming aminoacyl adenylate and releasing pyrophosphate.

This reaction is catalysed by a large number of enzymes, but each is specific for a particular amino acid. This means at least twenty two enzymes must function at this stage. These enzymes are generally known as amino acid activating enzyme. Magnesium ion (as well as ATP) is involved in this reaction.

Then each enzyme bound aminoacyl adenylate reacts immediately with tRNA and form aminoacyl-tRNA product. The same enzyme that is involved in activation also functions to transfer the amino acid to tRNA. These enzymes are then called aminoacyl-tRNA synthetase. They are unique catalysts, as they not only activate the amino acid but are also capable of recognizing a particular tRNA molecule.

During the formation of aminoacyl-tRNA product, the amino acid is transferred to the adenine that is present at one end of the tRNA molecules. When aminoacyl-tRNA product is formed, adenosine monophosphate and the aminoacyl synthetase are released. The tRNA functions as an adapter accepting amino acid and there is a group of three bases called anticodon. The anticodon is complementary with a triplet codon present in mRNA.

2. Transfer reaction:

Perhaps the most important step in protein synthesis is the transfer reaction. It involves aminoacyl-tRNA, ribosomes, and mRNA resulting in peptide-bond formation. For this reaction two enzymes are required as well as guanosine triphosphate (GTP) and several inorganic ions.

The ribosomes are not active until they are first combined with mRNA. A small ribosomal subunit become attached to the mRNA near initiation codon (AUG). The first step in transfer reaction is the binding reaction which occurs between aminoacyl- tRNA and the mRNA ribosome complex. It appears to be non enzymatic process, however, the binding is very specific.

That is an aminoacyl-tRNA carrying a specific anticodon fits in a proper position of the mRNA containing the corresponding codon. Initiator tRNA (with format) pairs with this codon and then a large ribosomal subunit joins to the small sub unit to initiate translation.

Magnesium ions are required for the stable bonding of mRNA to tRNA and also for the stability of the ribosomes. Each ribosome contains two sites, the P (for polypeptide) site and the A (for amino acid) site. A t-RNA with attached polypeptide is at the P-site and a t-RNA amino acid complex just arrives at the A- site. Further the attachment of the ribosome to the messenger is also quite specific and always occurs in a single site on the 5′- hydroxyl end of the mRNA.

The mRNA in a linear form passes through or over the ribosome. At the ribosome surface the complementary tRNAs with amino acids line up along the mRNA. The amino acid of each tRNA molecule interacts with the preceding amino acid forming a peptide link and releasing the preceding tRNA molecule. The enzyme peptidyl transferase and the energy rich molecule GTP are involved in the process.

As the mRNA moves through the ribosome from a site to the empty -P site and becomes free, again another ribosome may attach itself to the beginning of the molecule. Since the ribosome has moved forward three nucleotides, there is a new codon now located at the empty A-site.

Electron microscopic picture show many ribosomes attached at intervals along the length of a mRNA molecule. This complex is known as Polysome. When a peptide chain is completed, it is released and becomes a protein molecule. All the released tRNA molecules are once again charged with amino acids and the transfer reaction is repeated again and again.

The synthesis of protein chain begins at the amino end of polypeptide chain and progresses to the carboxyl end at the rate of two amino acids per second. Termination of polypeptide synthesis occurs at a stop codon i.e., UAA, UGA and UAG which does not code for any amino acid.

The arrival of stop codon in A- site results in the completion of protein synthesis and enzymatic cleavage of last tRNA from polypeptide. Although we know so much about protein synthesis, we still need to know which of the two strands of DNA, double helix acts as a template to form mRNA.