polypeptide. Of the three steps - initiation, elongation and termination, the first is the rate-limiting and the most highly regulated one. Along the route, three tRNA sites are present: the A, P and E sites for aminoacyl-tRNA (aa-tRNA), peptidyl-tRNA and deacylated-tRNA binding, respectively. First, an initiator methionyl tRNA (Met-tRNA) in a ternary complex (TC) with initiation factor and GTP binds the small 40S ribosomal subunit aided by several factors and forming a preinitiation complex (PIC). Binding to an activated 5'-capped mRNA prompts scanning of mRNA sequence. Once the initiating AUG codon is found, the formation of a scanning-arrested complex is prompted, followed by 60S subunit joining and formation of elongation competent 80S ribosome. During elongation, elongating aa-tRNAs in ternary complex with elongation factor and GTP are rapidly sampled until a cognate tRNA is found. Cognate tRNA binding, GTP hydrolysis, factor dissociation and peptidyl transfer are followed by the translocation of the elongated tRNA from A to P site and of deacylated-tRNA from P to E site, to complete one elongation cycle. Reading of a stop codon leads to translation termination followed by splitting of the two ribosomal subunits and their recycling. Recycling also occurs due to aberrantly stalled ribosomes. Control/surveillance mechanisms are in place to assure the rescue of stalled ribosomes due to structural features of mRNA, the presence of a premature stop codon or the absence of one. As a whole, translation is conserved but differences exist between the kingdoms of life. Eukaryotic translation has best been studied in prokaryotes and yeast, but evidence from mammalian systems is accumulating. Translation as currently known in higher eukaryotes is presented here.

Translation initiation pathway
Initiation starts with the formation of a ternary complex (TC) between initiation factor Eif2, initiator methionyl-tRNA (Met-tRNA) and GTP. Eif2 is a trimer whose alpha and beta subunits bind the gamma subunit which interacts with both GTP and tRNA. Binding of TC to the 40S decoding subunit is facilitated by factors Eif1, 1a, 3 and 5 forming a 43S preinitiation complex (PIC). Almost as large as 40S, Eif3 is a multisubunit complex that spans the solvent side of 40S and may serve to coordinate PIC assembly. Eif1 and 1a bind near P and A tRNA binding sites, respectively; their cooperative binding is thought to promote an 'open' PIC conformation poised to assemble on mRNA and scan it.
Mature mRNAs have a 7-methyl-guanine cap (m7G) on the 5'-end a polyadenylated tail (poly(a)) on the 3'-end. mRNAs are 'primed' for 43S recruitment by a set of factors that include the EIF4F complex and polyA binding protein Pabp. EIF4F consists of Eif4e cap binding factor, the RNA helicase Eif4a and the Eif4g scaffold which binds both factors and mRNA. The helicase activity of Eif4a is stimulated by Eif4b and 4h; RNA unwinding and removal of secondary structures prepare PIC binding which requires single-stranded mRNA. Stable mRNA structures may require the activity of additional helicases and accompanying ATP hydrolysis to allow scanning; such is the case for Dhx29, for instance. The interaction of Eif4g with both Eif4e and Pabp circularizes mRNA and the 'closed-loop' conformation promotes PIC recruitment; however, this appears to vary with cell type. The exact mechanism of mRNA-PIC recruitment remains unclear; in mammals, Eif4g-Eif3 interaction plays a central role. Once at 5' of mRNA, PIC needs to travel in the 5'-3' direction to find the initiation codon, which can be located far downstream. It is believed that it proceeds linearly, on a base-by-base scanning. AUG finding and complementarity with the anticodon of initiator Met-tRNA in the P site, triggers hydrolysis of GTP by Eif2, likely aided by the GAP (GTPase activating protein) activity of Eif5. However, it is only after ejection of Eif1 from PIC and release of the gating inorganic phosphorus (Pi) by Eif2 that PIC will undergo the transition to the 'closed', scanning-arrested conformation of 48S. With the closing of PIC, Eif2-GDP and Eif5 must dissociate and the 60S subunit has to join in to form the 80S IC - a process facilitated by Eif5b GTPase. Eif5b interaction with Eif1a is important for this step; Eif5b has binding sites on 80S IC as well as contact sites on both ribosomal subunits. Hydrolysis of its own bound GTP is not necessary for 60S joining but triggers its release from 80S IC. Finally, release of Eif1a renders 80S IC in position to start elongation. Eif2-GDP is recycled to Eif2-GTP by the heteropentameric Eif2b complex.

Translation elongation pathway
At the beginning of an elongation step, the P site is occupied by peptidyl-tRNA (Met-tRNA at the end of the initiation), the E site by a deacylated-tRNA (empty at the end of initiation), and the A site is available for loading aa-tRNA. aa-tRNAs are delivered to the A site decoding center in a ternary complex (TC) with elongation factor Tu (Eef1a1) and GTP. Due to genetic code degeneracy, a single amino acid may have more than one isoacceptor tRNA. The overall tRNA shape has a cloverleaf secondary, and L-shaped tertiary structure. The anticodon stem loop (ASL) is at one extremity, and the attached amino acid is at the 3'-end CCA sequence. Elongation is the most evolutionarily conserved step; its proceedings, mostly investigated in the prokaryotic system, occur similarly in eukaryotes. Initial sampling of TCs is rapid until a match is found and base-pairing of cognate tRNA anticodon and mRNA codon at the A site occurs. Conformational changes in the small subunit lead to rRNA interactions with the codon-anticodon helix and domain 'closure'. Additional structural changes occur in tRNA and the elongation factor. Together, they promote Eef1a1 activation and GTP hydrolysis; Pi release and conformational changes in the GDP-bound factor lead to its dissociation from the ribosome. The now free 3'-acceptor end of aa-tRNA moves into the peptidyl transfer center; peptide bond formation proceeds via a nucleophilic attack of the alpha-amino group of the aa-tRNA of A site on the ester bond of peptidyl-tRNA at P site, a reaction enhanced by the ribosome. The 3'-acceptor end of now deacylated tRNA at the P site moves into the E site. At this point, the ribosome is in the PRE-translocation configuration with hybrid A/P and P/E tRNA states; the first letter indicates location on the small subunit and the second the one on the large. Translocation of tRNAs - the movement of codon-anticodon duplexes on the small subunit from A and P sites to P and E sites, respectively and of the next codon into the A site, is catalyzed by elongation factor Eef2. Eef2-GTP binding the PRE-ribosome induces a conformational change whereby the small subunit is rotated with respect to the large one. GTP hydrolysis precedes and accelerates translocation; the ribosome is now in the POST state with classical A/A, P/P and E/E sites. The deacylated tRNAs leaving the ribosome replenish the pool of substrates for the family of aminoacyl-tRNA synthetases, and therefore of TCs available for elongation - evidence suggests that the flow of tRNA during elongation follows a closed circuit, referred to as the tRNA cycle. Eef1a1 is cycled back to its GTP-form by Eef1b2.

Translation termination pathway
Termination of translation occurs when a stop codon - UAG, UAA or UGA, enters the A site. Two release factors participate in this step: the class I factor Etf1 (known as Erf1) and the class II factor Gspt1 (known as Erf3). Etf1 is involved in codon recognition and peptidyl-tRNA hydrolysis; Gspt1 is a GTPase aiding in the process. GTP hydrolysis promotes proper positioning of the M domain of Etf1 into the P site and accelerates peptide release. The middle (M) domain of Etf1 contains a universally conserved Gly-Gly-Gln (GGQ) motif that is essential for hydrolysis while the N- and C- termini are involved in codon recognition and interaction with Gspt1, respectively. The mechanism of peptide release resembles the one of peptide bond formation. The distinction lies in the identity of the nucleophile - here, water is the nucleophile attacking the ester bond of the peptidyl-tRNA at P site, and the requirement for protein factors in peptide release. Dissociation and subsequent recycling of ribosomal subunits is mediated by Abce1. Abce1 is the only member of subfamily E of the ATP binding cassette (ABC) family of ATPases and unique in possessing iron-sulfur (FeS) cluster domain at its N-terminus. Mechanistically is not clear how Abce1 promotes dissociation of ribosomal subunits but it is thought that ATP hydrolysis provides the energy. Ribosome recycling is considered a fourth step of translation. Ribosome recycling/rescue is also prompted when aberrant stalling occurs. Ribosome can stall due to mRNA features such as pseudoknots, rare codons, stem loops and others, the absence of a stop codon or its premature presence. In these cases, the mRNA surveillance pathways of no-go decay (NGD), nonstop decay (NSD) and nonsense-mediated mRNA decay (NMD) are triggered, respectively. In NMD, Etf1 and Gspt1 recognize the stalled ribosomes; in NGD and NSD, they are recognized by the two corresponding paralogs - Pelota (Pelo) and Hbs1, respectively. Abce1 interacts with Etf1 or Pelo to promote recycling.

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