translation (genetics) - Prokaryotic translation, Eukaryotic translation, Translation by hand, Translation by computer

The making of a protein from a messenger RNA which takes place on a ribosome in the cytoplasm of the cell. Each three nucleotide bases of the mRNA represent a specific amino acid which are joined together in succession.

Translation is the second process of protein biosynthesis (part of the overall process of gene expression).Translation occurs in the cytoplasm where the ribosomes are located. In translation, messenger RNA (mRNA) is decoded to produce a specific polypeptide according to the rules specified by the genetic code. Translation proceeds in four phases: activation, initiation, elongation and termination (all describing the growth of the amino acid chain, or polypeptide that is the product of translation). While this is not technically a step in translation, it is required for translation to proceed. Initiation involves the small subunit of the ribosome binding to 5' end of mRNA with the help of initiation factors (IF), other proteins that assist the process. Termination of the polypeptide happens when the A site of the ribosome faces a stop (nonsense) codon (UAA, UAG, or UGA). The ribosome and tRNA molecules translate this code to produce proteins. tRNAs have a site for amino acid attachment, and a site called an anticodon. This aminoacyl-tRNA travels inside the ribosome, where mRNA codons are matched through complementary base pairing to specific tRNA anticodons.

Prokaryotic translation

Initiation

Initiation of translation in prokaryotes involves the assembly of the components of the translation system which are: the two ribosomal subunits, the mRNA to be translated, the first aminoacyl tRNA (the tRNA charged with the first amino acid), GTP (as a source of energy), and initiation factors which help the assembly of the initiation complex. Prokaryotic initiation results in the association of the small and large ribosomal subunits and binding of first aminoacyl tRNA (fmet-tRNA) through anticodon-codon base pairing with the initiation codon of mRNA.

The ribosome consists of three sites: the A site, the P site, and the E site.

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. The 16s rRNA of the small 30S ribosomal subunit recognizes the ribosomal binding site on mRNA (the Shine-Dalgarno sequence, 5-10 base pairs upstream of the start codon(AUG)) The Shine-Delgarno 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. IF-3 helps to position fmet-tRNA into the P site, such that fmet-tRNA interacts via base pairing with the mRNA initiation codon (AUG). Note that prokaryotes can differentiate between a normal AUG (coding for methionine) and an AUG initiation codon (coding for formylmethionine and indicating the start of a new translation process).

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. Now the P site contains the beginning of the peptide chain of the protein to be encoded and the A site has the next base 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. Now, the A site has newly formed peptide, while the P site has an unloaded tRNA (tRNA with no amino acids). 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

Termination occurs when one of the three termination codons moves into the A site.

Recycling

The post-termination complex formed by the end of the termination step consists of mRNA with the termination codon at the A-site, tRNAs and the ribosome. 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 ribosomes into the 30s and 50s subunits. This "recycles" the ribosomes for additional rounds of translation.

Polysomes

Translation is carried out by more than one ribosome simultaneously. Tetracyclines block the A site on the ribosome, preventing the binding of aminoacyl tRNAs.

Eukaryotic translation

Initiation

The cap-dependent initiation

Initiation of translation involves an interaction of some proteins with a special tag bound to 5'-end of the mRNA molecules. The subunit accompanied by some of those protein factors moves along the mRNA chain towards its 3'-end and scans for the 'start' codon (mostly AUG) on the mRNA, which indicates where the mRNA starts coding for the protein. The initiator tRNA charged with Met forms part of the ribosomal complex and thus all proteins start with this amino acid (unless it is cleaved away by a protease in some subsequent steps).

The cap-independent initiation

The best studied example of the cap-independent mode of translation initiation in eukaryotes is the Internal Ribosome Entry Site IRES approach. What differentiates cap-independent translation from cap-dependent translation is that cap-independent translation does not require the ribosome to start scanning from the 5' end of the mRNA cap until the start codon. The ribosome can be trafficked to the start site by ITAFs (IRES trans-acting factors) bypassing the need to scan from the 5' end of the untranslated region of the mRNA. This method of translation has been recently discovered, and has found to be important in conditions that require the translation of specific mRNAs, despite cellular stress or the inability to translate most mRNAs.

cap-dependent/cap-independent initiation applies to both prokaryotes and eukaryotes.

Translation by hand

It is also possible to translate either by hand (for short sequences) or by computer (after first programming one appropriately, see section below), this allows biologists and chemists to draw out the chemical structure of the encoded protein on paper. Note that there are 3 translation "windows" depending on where you start reading the code.

This approach may not give the correct amino acid composition of the protein, in particular if unconventional amino acids such as selenocysteine are incorporated into the protein, which is coded for by a conventional stop codon in combination with a downstream hairpin (SElenoCysteine Insertion Sequence, or SECIS).

Translation by computer

An enormous amount of computer programs capable of translating a DNA/RNA sequence into protein sequence exist. However, few programs can handle all the "special" cases, such as the use of the alternative initiation codons: For examle the rare alternative start codon TTG codes for Methionine when used as a start codon and for Leucine in all other positions.

Example: Condensed translation table for the Standard Genetic Code (from the NCBI Taxonomy webpage).

AAs = FFLLSSSSYY**CC*WLLLLPPPPHHQQRRRRIIIMTTTTNNKKSSRRVVVVAAAADDEEGGGG Starts = ---M---------------M---------------M---------------------------- Base1 = TTTTTTTTTTTTTTTTCCCCCCCCCCCCCCCCAAAAAAAAAAAAAAAAGGGGGGGGGGGGGGGG Base2 = TTTTCCCCAAAAGGGGTTTTCCCCAAAAGGGGTTTTCCCCAAAAGGGGTTTTCCCCAAAAGGGG Base3 = TCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAGTCAG

Translation tables

Even when working with ordinary Eukaryotic sequences such as the Yeast genome, it is often desired to be able to use alternative translation tables -- namely for translation of the mitochondrial genes. Currently the following translation tables are defined by the NCBI Taxonomy Group for the translation of the sequences in GenBank:

1: The Standard Code 2: The Vertebrate Mitochondrial Code 3: The Yeast Mitochondrial Code 4: The Mold, Protozoan, and Coelenterate Mitochondrial Code and the Mycoplasma/Spiroplasma Code 5: The Invertebrate Mitochondrial Code 6: The Ciliate, Dasycladacean and Hexamita Nuclear Code 9: The Echinoderm and Flatworm Mitochondrial Code 10: The Euplotid Nuclear Code 11: The Bacterial and Plant Plastid Code 12: The Alternative Yeast Nuclear Code 13: The Ascidian Mitochondrial Code 14: The Alternative Flatworm Mitochondrial Code 15: Blepharisma Nuclear Code 16: Chlorophycean Mitochondrial Code 21: Trematode Mitochondrial Code 22: Scenedesmus obliquus mitochondrial Code 23: Thraustochytrium Mitochondrial Code

Software examples

ApE (Mac, Windows, Unix) DNA Strider (Mac) ExPASy Translate Tool (webserver) Virtual Ribosome (webserver, cross-platform command-line)

Example of computational translation - notice the indication of (alternative) start-codons:

VIRTUAL RIBOSOME ---------------- Translation table: Standard SGC0 >Seq1 Reading frame: 1 M V L S A A D K G N V K A A W G K V G G H A A E Y G A E A L 5' ATGGTGCTGTCTGCCGCCGACAAGGGCAATGTCAAGGCCGCCTGGGGCAAGGTTGGCGGCCACGCTGCAGAGTATGGCGCAGAGGCCCTG 90 >>>...)))..............................................................................))) E R M F L S F P T T K T Y F P H F D L S H G S A Q V K G H G 5' GAGAGGATGTTCCTGAGCTTCCCCACCACCAAGACCTACTTCCCCCACTTCGACCTGAGCCACGGCTCCGCGCAGGTCAAGGGCCACGGC 180 ......>>>...))).......................................)))................................. A K V A A A L T K A V E H L D D L P G A L S E L S D L H A H 5' GCGAAGGTGGCCGCCGCGCTGACCAAAGCGGTGGAACACCTGGACGACCTGCCCGGTGCCCTGTCTGAACTGAGTGACCTGCACGCTCAC 270 ..................)))..................)))......))).........)))......)))......)))......... K L R V D P V N F K L L S H S L L V T L A S H L P S D F T P 5' AAGCTGCGTGTGGACCCGGTCAACTTCAAGCTTCTGAGCCACTCCCTGCTGGTGACCCTGGCCTCCCACCTCCCCAGTGATTTCACCCCC 360 ...)))...........................))).........))))))......)))..............................