The ribosome is a cellular machine found in all organisms. It serves to convert the instructions found in messenger RNA (mRNA, which itself is made from instructions in DNA) into the chains of amino acids that make up proteins. It is a sphere-shaped structure within the cytoplasm of a cell that is composed of RNA and protein. It is the site of protein synthesis. Ribosomes are free in the cytoplasm and often attached to the membrane of the endoplasmic reticulum. Ribosomes exist in both eukaryotic and prokaryotic cells. Plastids and mitochondria, in eukaryotic cells, have smaller ribosomes similar to those of prokaryotes.
Ribosomes are ribonucleoprotein particles that contain rRNA and r-proteins. It acts like a small migrating factory. Each ribosome is made up of two subunits – large and small subunits. Each of the ribosome subunits contains rRNA and a number of small proteins. In E.coli, the 30S subunit consists 16S rRNA (1541 nucleotides) and 21 r – proteins. The 50S subunit contains 23S rRNA (2904 nucleotides), the small 5S RNA (120 nucleotides) and 31 proteins. The cytosolic ribosomes of eukaryotes are larger than those of bacteria. The total content of both RNA and protein is greater; the major RNA molecules are longer (called 18S and 28S rRNAs) and there are more proteins. The 60S subunit contains 28S rRNA (4718 nucleotides), the small 5S rRNA (120 nucleotides), 5.8S rRNA (160 nucleotides) and ~50 proteins. The 40S subunit consists of the 18S rRNA (1874 nucleotides) and 33 r-proteins.
Location in the Cell
There are two places where ribosomes usually exist in the cell; it is either suspended in the cytosol and/or bound to the endoplasmic reticulum. These ribosomes are called free ribosomes which are bound ribosomes respectively. In both cases, the ribosomes usually form aggregates called polysomes or polyribosomes during protein synthesis. Free ribosomes usually make proteins that will function in the cytosol (fluid component of the cytoplasm), while bound ribosomes usually make proteins that are exported from the cell or included in the cell’s membrane. Interestingly enough, free ribosomes and bound ribosomes are interchangeable and the cell can change their numbers according to metabolic needs.
The RNA binding Sites in the Ribosome: The 70S ribosome has 3 tRNA-binding sites
- P-site also called the peptidyl-tRNA-binding site holds the tRNA molecule that is linked to the growing end of the polypeptide chain.
- A-site also called the aminoacyl-tRNA binding site holds the incoming tRNA molecule charged with an amino acid.
- E-site Deacylated tRNA (lacking any amino acid) exits via the E site, also called the exit site.
When the initiator tRNA joins the 30S subunit of the prokaryotic ribosome with its mRNA attached, it fits into one of the three sites in the ribosome. These sites in the ribosome are referred to as aminoacyl site (A site), peptidyl site (P site) and exit site (E site). A and P sites each contain a tRNA just before forming a peptide bond: the P site contains the tRNA with the growing peptide chain; the A site contains a new tRNA with its single amino acid. The exit site helps eject depleted tRNAs after a peptide bond forms.
Amino acid activation and attachment to tRNA
The attachment of amino acids to tRNAs is the function of the group of enzymes called aminoacyl-tRNA synthetases. Aminoacyl-tRNA synthetases activate the amino acids by covalently linking them to tRNAs. When a tRNA is charged with the amino acid corresponding to its anticodon, it is called aminoacyl-tRNA. Aminoacyl-tRNA synthetases catalyze this reaction in two steps. In the first step, an activated amino acid intermediate is formed by reaction between the amino acid and ATP and in the second step the amino acid is transferred to the 3ˈ end of the tRNA, the link is formed between the –COOH group of the amino acid and the -OH group attached to either the 2ˈ or 3ˈ carbon on the sugar of the last nucleotide, which is always an A. In aminoacylation, the amino acid is linked to the tRNA by a high energy bond and thus is said to be activated
Challenges faced by aminoacyl tRNA synthetases in selecting the correct amino acid are more daunting than its recognition of the appropriate tRNA. It has a catalytic pocket for adenylation and editing pocket to allow the tRNA to proofread the product of the adenylation reaction.
With a few exceptions, organisms have 20 aminoacyl-tRNA synthetases, one for each amino acid. Attachment of the appropriate amino acid to a tRNA is catalyzed by a specific aminoacyl-tRNA synthetase. Each of the 20 different synthetases recognizes one amino acid and all its compatible or cognate, tRNAs. Synthetases recognize the anticodon loops and acceptor stems of transfer RNA molecules. These coupling enzymes link an amino acid to the free 2ˈ or 3ˈhydroxyl of the adenosine at the 3ˈ terminus of tRNA molecules by an ATP-requiring reaction. The 20 aminoacyl-tRNA synthetases fall into two distinct groups – Class I and Class II. Class I enzyme attach the amino acid to the 2ˈ-OH group of the terminal nucleotide of the tRNA, whereas Class II enzymes attach the amino acid to the 3ˈ-OH group. When an amino acid has become attached to a tRNA molecule, the tRNA is said to be acylated or charged. The term uncharged tRNA refers to a tRNA molecule lacking an amino acid and mischarged tRNA to one acylated with an incorrect amino acid.