The ribosome consists of three sites: the A site, the P site, and the E site. The A site is the point of entry for the aminoacyl tRNA (except for the first aminoacyl tRNA, fMet-tRNAfMet, which enters at the P site). The P site is where the peptidyl tRNA is formed in the ribosome. And the E site which is the exit site of the now uncharged tRNA after it gives its amino acid to the growing peptide chain.
Initiation of translation begins with the 50S and 30S ribosomal subunits dissociated. IF1 (initiation factor 1) blocks the A site to insure that the fMet-tRNA can bind only to the P site and that no other aminoacyl-tRNA can bind in the A site during initiation, while IF3 blocks the E site and prevents the two subunits from associating. IF2 is a small GTPase which binds fmet-tRNAfMet and helps its binding with the small ribosomal subunit. The 3' end of the 16S rRNA of the small 30S ribosomal subunit recognizes the ribosomal binding site on the mRNA (Shine-Dalgarno sequence or SD), through its anti-SD sequence, 5-10 base pairs upstream of the start codon. The Shine-Dalgarno sequence is found only in prokaryotes. This helps to correctly position the ribosome onto the mRNA so that the P site is directly on the AUG initiation codon. IF3 helps to position fMet-tRNAfMet into the P site, such that fMet-tRNAfMet interacts via base pairing with the mRNA initiation codon (AUG). Initiation ends as the large ribosomal subunit joins the complex causing the dissociation of initiation factors. Note that prokaryotes can differentiate between a normal AUG (coding for methionine) and an AUG initiation codon (coding for formyl-methionine and indicating the start of a new translation process).
Elongation of the polypeptide chain involves addition of amino acids to the carboxyl end of the growing chain. The growing protein exits the ribosome through the polypeptide exit tunnel in the large subunit.
Elongation starts when the fmet-tRNA enters the P site, causing a conformational change which opens the A site for the new aminoacyl-tRNA to bind. This binding is facilitated by elongation factor-Tu (EF-Tu), a small GTPase. Now the P site contains the beginning of the peptide chain of the protein to be encoded and the A site has the next amino acid to be added to the peptide chain. The growing polypeptide connected to the tRNA in the P site is detached from the tRNA in the P site and a peptide bond is formed between the last amino acids of the polypeptide and the amino acid still attached to the tRNA in the A site. This process, known as peptide bond formation, is catalyzed by a ribozyme, peptidyltransferase, an activity intrinsic to the 23S ribosomal RNA in the 50S ribosomal subunit. Now, the A site has newly formed peptide, while the P site has an unloaded tRNA (tRNA with no amino acids). In the final stage of elongation, translocation, the ribosome moves 3 nucleotides towards the 3'end of mRNA. Since tRNAs are linked to mRNA by codon-anticodon base-pairing, tRNAs move relative to the ribosome taking the nascent polypeptide from the A site to the P site and moving the uncharged tRNA to the E exit site. This process is catalyzed by elongation factor G (EF-G).
The ribosome continues to translate the remaining codons on the mRNA as more aminoacyl-tRNA bind to the A site, until the ribosome reaches a stop codon on mRNA(UAA, UGA, or UAG).
Termination occurs when one of the three termination codons moves into the A site. These codons are not recognized by any tRNAs. Instead, they are recognized by proteins called release factors, namely RF1 (recognizing the UAA and UAG stop codons) or RF2 (recognizing the UAA and UGA stop codons). These factors trigger the hydrolysis of the ester bond in peptidyl-tRNA and the release of the newly synthesized protein from the ribosome. A third release factor RF-3 catalyzes the release of RF-1 and RF-2 at the end of the termination process.
The post-termination complex formed by the end of the termination step consists of mRNA with the termination codon at the A-site, an uncharged tRNA in the P site, and the intact 70S ribosome. Ribosome recycling step is responsible for the disassembly of the post-termination ribosomal complex. Once the nascent protein is released in termination, Ribosome Recycling Factor and Elongation Factor G (EF-G) function to release mRNA and tRNAs from ribosomes and dissociate the 70S ribosome into the 30S and 50S subunits. IF3 then replaces the deacylated tRNA releasing the mRNA. All translational components are now free for additional rounds of translation.
Studies from Pohang University of Science and Technology (POSTECH) Have Provided New Information about Metabolic Engineering
Feb 06, 2013; Research findings on Metabolic Engineering are discussed in a new report. According to news originating from Gyeongbuk, South...