Release factor

Classification

There are two classes of release factors. Class 1 release factors recognize stop codons; they bind to the A site of the ribosome in a way mimicking that of tRNA, releasing the new polypeptide as it disassembles the ribosome. Class 2 release factors are GTPases that enhance the activity of class 1 release factors. It helps the class 1 RF dissociate from the ribosome.

Bacterial release factors include RF1, RF2, and RF3 (or PrfA, PrfB, PrfC in the "peptide release factor" gene nomenclature). RF1 and RF2 are class 1 RFs: RF1 recognizes UAA and UAG while RF2 recognizes UAA and UGA. RF3 is the class 2 release factor. Eukaryotic and archaeal release factors are named analogously, with the naming changed to "eRF" for "eukaryotic release factor" and vice versa. a/eRF1 can recognize all three stop codons, while eRF3 (archaea use aEF-1α instead) works just like RF3.

The bacterial and archaeo-eukaryotic release factors are believed to have evolved separately. The two groups class 1 factors do not show sequence or structural homology with each other. The homology in class 2 is restricted to the fact that both are GTPases. It is believed that (b)RF3 evolved from EF-G while eRF3 evolved from eEF1α.

In line with their symbiotic origin, eukaryotic mitochondria and plastids use bacterial-type class I release factors. As of April 2019, no definite reports of an organellar class II release factor can be found.

Human genes

• RF1 (mitochondrial): MTRF1, MTRF1L, MRPL58 (ICT1), MTRFR (C12orf65)
• eRF1: ETF1
• eRF3: GSPT1, GSPT2

Structure and function

Crystal structures have been solved for bacterial 70S ribosome bound to each of the three release factors, revealing details in codon recognition by RF1/2 and the EF-G-like rotation of RF3. Cryo-EM structures have been obtained for eukaryotic mamallian 80S ribosome bound to eRF1 and/or eRF3, providing a view of structural rearrangements caused by the factors. Fitting the EM images to previously known crystal structures of individual parts provides identification and a more detailed view of the process.

In both systems, the class II (e)RF3 binds to the universal GTPase site on the ribosome, while the class I RFs occupy the A site.

Bacterial

The bacterial class 1 release factors can be divided into four domains. The catalytically-important domains are:

• The "tripeptide anticodon" motif in domain 2, P[AV]T in RF1 and SPF in RF2. Only one residue actually participates in stop codon recognition via hydrogen bonding.
• The GGQ motif in domain 3, critical for peptidyl-tRNA hydrolase (PTH) activity.

As RF1/2 sits in the A site of the ribosome, domains 2, 3, and 4 occupy the space that tRNAs load into during elongation. Stop codon recognition activates the RF, promoting a compact to open conformation change, sending the GGQ motif to the peptidyl transferase center (PTC) next to the 3′ end of the P-site tRNA. By hydrolysis of the peptidyl-tRNA ester bond, which displayed pH-dependence in vitro, the peptide is cut loose and released. RF3 is still needed to release RF1/2 from this translation termination complex.

After releasing the peptide, ribosomal recycling is still required to empty the P-site tRNA and mRNA out to make the ribosome usable again. This is done by splitting the ribosome with factors like IF1–IF3 or RRF–EF-G.

Eukaryotic and archaeal

eRF1 can be broken down into four domains: N-terminal (N), Middle (M), C-terminal (C), plus a minidomain:

• The N domain is responsible for stop codon recognition. Motifs include TASNIKS and YxCxxxF.
• A GGQ motif in the M domain is critical for peptidyl-tRNA hydrolase (PTH) activity.

Unlike in the bacterial version, eRF1–eRF3–GTP binds together into a sub-complex, via a GRFTLRD motif on RF3. Stop codon recognition makes eRF3 hydrolyze the GTP, and the resulting movement puts the GGQ into the PTC to allow for hydrolysis. The movement also causes a +2-nt movement of the toeprint of the pre-termination complex. The archaeal aRF1–EF1α–GTP complex is similar. The triggering mechanism is similar to that of aa-tRNA–EF-Tu–GTP.

A homologous system is Dom34/Pelota–Hbs1, a eukaryotic system that breaks up stalled ribosomes. It does not have GGQ. The recycling and breakup is mediated by ABCE1.