Translation occurs using ribosomes, which are present in the cytosol of eukaryotic cells. mRNA of eukaryotic cells has to leave the nucleus and move to the cytoplasm for translation to occur. In prokaryotic cells, ribosomes are capable of attaching to the mRNA while it is still being translated from DNA.This is possible since the translation begins at the 5' end of the mRNA and transcription produces mRNA in a 5' to 3' direction, so the 5' end is created first. Since transcription and translation are done simultaneously, mRNA is more short-lived compared to that of eukaryotes. The multiple steps involved with the translation of mRNA in eukaryotic cells allow for greater ability to regulate protein production.
Ribosomes are composed of two subunits, a large one and a small one, that assemble at the mRNA and otherwise exist separately in the cytoplasm. The ribosomes contain proteins, ribosomal RNA (rRNA), and transfer RNA (tRNA). tRNA are adaptor molecules with a 3' end and an anticodon. The anticodon reads the triplet codon of the mRNA using complementary base pairing and the 3' end is attached to a particular amino acid. rRNA catalyzes the addition of amino acids to the growing protein.
There is an area of the mRNA near the 5' end that is not translated, and is known as the UTR or untranslated region. The UTR is located between the first nucleotide and the start codon of the mRNA strand. The UTR is important to translation because it provides a binding site for the ribosome. The human mRNA UTR is typically about 170 nucleotides long. Translation begins after a complex is formed on the mRNA strand. First, three initiation factor proteins bind to the smaller ribosomal subunit, after which this complex along with a tRNA that contains methionine bind to the mRNA that is being translated, close to the AUG start codon. While methionine is the first amino acid of all new proteins, it can be removed after translation. The second amino acid on the protein influences whether or not an enzyme removes the initial methionine. For example, if the amino acid alanine is the second amino acid, then the initial methionine is removed by an enzme. If the second amino acid is lysine, then the methionine is not removed.
The initiation factors are removed from the initiation complex on the mRNA when the large ribosomal subunit binds to the complex. This subunit contains three sites that the tRNA can bind to: an A site, an E site, and a P site. The mRNA codon and the aminoacyl-tRNA anticodon pair up at the A site to ensure that the correct amino acid is added to the polypeptide. The P site is where the amino acid on the 3' end of the tRNA is transferred to the polypeptide chain. The E site is where the "empty" tRNA stays before it is released into the cytoplasm. Only a tRNA containing methionine is able to bind to the ribosome's P site while the second mRNA codon lines up at the A site.
The ribosome moves along the mRNA in a 5' to 3' direction and covalently binds amino acids to one another, producing a polypeptide. This process requires GTP as an energy source and elongation factors. When the tRNA-amino acid complex binds to the A site, GTP is cleaved to form GDP and is released with certain elongation factors and is later recycled by other elongation factors. Peptidyl transferase activity binds the first and second amino acids together. This is a catayltic function of rRNA, although this process was originally thought to have been catalyzed by enzymes. Once the bond is formed, the ribosome shifts so that the now-empty tRNA molecule is in the E site. The tRNA is then released by the ribosome into the cytoplasm, where it picks up another amino acid to repeat the process. The A site is also empty for the next tRNA molecule that corresponds to the next codon. The protein-coding sequence of mRNA ends with UAA, UAG, and UGA. tRNAs do not recognize these codons and another protein, called a release factor, binds to it and causes the polypeptide to be removed from the ribosome and the ribosome disassociates.
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