Release factor

Last updated
Peptide chain release factor, bacterial Class 1
Identifiers
SymbolPCRF
Pfam PF03462
InterPro IPR005139
Available protein structures:
Pfam   structures / ECOD  
PDB RCSB PDB; PDBe; PDBj
PDBsum structure summary
Peptide chain release factor, bacterial Class 1, PTH domain, GGQ
Identifiers
SymbolRF-1
Pfam PF00472
Pfam clan CL0337
InterPro IPR000352
PROSITE PS00745
Available protein structures:
Pfam   structures / ECOD  
PDB RCSB PDB; PDBe; PDBj
PDBsum structure summary
Peptide chain release factor eRF1/aRF1
Identifiers
Symbol?
InterPro IPR004403

A release factor is a protein that allows for the termination of translation by recognizing the termination codon or stop codon in an mRNA sequence. They are named so because they release new peptides from the ribosome.

Contents

Background

During translation of mRNA, most codons are recognized by "charged" tRNA molecules, called aminoacyl-tRNAs because they are adhered to specific amino acids corresponding to each tRNA's anticodon. In the standard genetic code, there are three mRNA stop codons: UAG ("amber"), UAA ("ochre"), and UGA ("opal" or "umber"). Although these stop codons are triplets just like ordinary codons, they are not decoded by tRNAs. It was discovered by Mario Capecchi in 1967 that, instead, tRNAs do not ordinarily recognize stop codons at all, and that what he named "release factor" was not a tRNA molecule but a protein. [1] Later, it was demonstrated that different release factors recognize different stop codons. [2]

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. [3] [4] 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. [5]

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. [6] 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. [6] [7]

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. [8] [9] 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. [10]

In line with their symbiotic origin, eukaryotic mitochondria and plastids use bacterial-type class I release factors. [11] As of April 2019, no definite reports of an organellar class II release factor can be found.

Human genes

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. [12] 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. [13] [14]

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. [12]

Bacterial

The bacterial class 1 release factors can be divided into four domains. The catalytically-import domains are: [12]

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, [15] 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, [16] the peptide is cut loose and released. RF3 is still needed to release RF1/2 from this translation termination complex. [12]

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. [17]

Eukaryotic and archaeal

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

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. [13] The archaeal aRF1–EF1α–GTP complex is similar. [18] The triggering mechanism is similar to that of aa-tRNA–EF-Tu–GTP. [14]

A homologous system is Dom34/Pelota–Hbs1, a eukaryotic system that breaks up stalled ribosomes. It does not have GGQ. [14] The recycling and breakup is mediated by ABCE1. [19] [20]

Related Research Articles

<span class="mw-page-title-main">Ribosome</span> Intracellular organelle consisting of RNA and protein functioning to synthesize proteins

Ribosomes ( ) are macromolecular machines, found within all cells, that perform biological protein synthesis. Ribosomes link amino acids together in the order specified by the codons of messenger RNA (mRNA) molecules to form polypeptide chains. Ribosomes consist of two major components: the small and large ribosomal subunits. Each subunit consists of one or more ribosomal RNA (rRNA) molecules and many ribosomal proteins. The ribosomes and associated molecules are also known as the translational apparatus.

<span class="mw-page-title-main">Stop codon</span> Codon that marks the end of a protein-coding sequence

In molecular biology, a stop codon is a codon that signals the termination of the translation process of the current protein. Most codons in messenger RNA correspond to the addition of an amino acid to a growing polypeptide chain, which may ultimately become a protein; stop codons signal the termination of this process by binding release factors, which cause the ribosomal subunits to disassociate, releasing the amino acid chain.

<span class="mw-page-title-main">Translation (biology)</span> Cellular process of protein synthesis

In molecular biology and genetics, translation is the process in which ribosomes in the cytoplasm or endoplasmic reticulum synthesize proteins after the process of transcription of DNA to RNA in the cell's nucleus. The entire process is called gene expression.

<span class="mw-page-title-main">Transfer RNA</span> RNA that facilitates the addition of amino acids to a new protein

Transfer RNA is an adaptor molecule composed of RNA, typically 76 to 90 nucleotides in length, that serves as the physical link between the mRNA and the amino acid sequence of proteins. tRNAs genes from Bacteria are typically shorter than tRNAs from Archaea and eukaryotes. The mature tRNA follows an opposite pattern with tRNAs from Bacteria being usually longer than tRNAs from Archaea, with eukaryotes exhibiting the shortest mature tRNAs. Transfer RNA (tRNA) does this by carrying an amino acid to the protein synthesizing machinery of a cell called the ribosome. Complementation of a 3-nucleotide codon in a messenger RNA (mRNA) by a 3-nucleotide anticodon of the tRNA results in protein synthesis based on the mRNA code. As such, tRNAs are a necessary component of translation, the biological synthesis of new proteins in accordance with the genetic code.

The Shine–Dalgarno (SD) sequence is a ribosomal binding site in bacterial and archaeal messenger RNA, generally located around 8 bases upstream of the start codon AUG. The RNA sequence helps recruit the ribosome to the messenger RNA (mRNA) to initiate protein synthesis by aligning the ribosome with the start codon. Once recruited, tRNA may add amino acids in sequence as dictated by the codons, moving downstream from the translational start site.

Ribosome display is a technique used to perform in vitro protein evolution to create proteins that can bind to a desired ligand. The process results in translated proteins that are associated with their mRNA progenitor which is used, as a complex, to bind to an immobilized ligand in a selection step. The mRNA-protein hybrids that bind well are then reverse transcribed to cDNA and their sequence amplified via PCR. The result is a nucleotide sequence that can be used to create tightly binding proteins.

Bacterial translation is the process by which messenger RNA is translated into proteins in bacteria.

Eukaryotic translation is the biological process by which messenger RNA is translated into proteins in eukaryotes. It consists of four phases: initiation, elongation, termination, and recapping.

<span class="mw-page-title-main">EF-Tu</span> Prokaryotic elongation factor

EF-Tu is a prokaryotic elongation factor responsible for catalyzing the binding of an aminoacyl-tRNA (aa-tRNA) to the ribosome. It is a G-protein, and facilitates the selection and binding of an aa-tRNA to the A-site of the ribosome. As a reflection of its crucial role in translation, EF-Tu is one of the most abundant and highly conserved proteins in prokaryotes. It is found in eukaryotic mitochondria as TUFM.

Eukaryotic initiation factors (eIFs) are proteins or protein complexes involved in the initiation phase of eukaryotic translation. These proteins help stabilize the formation of ribosomal preinitiation complexes around the start codon and are an important input for post-transcription gene regulation. Several initiation factors form a complex with the small 40S ribosomal subunit and Met-tRNAiMet called the 43S preinitiation complex. Additional factors of the eIF4F complex recruit the 43S PIC to the five-prime cap structure of the mRNA, from which the 43S particle scans 5'-->3' along the mRNA to reach an AUG start codon. Recognition of the start codon by the Met-tRNAiMet promotes gated phosphate and eIF1 release to form the 48S preinitiation complex, followed by large 60S ribosomal subunit recruitment to form the 80S ribosome. There exist many more eukaryotic initiation factors than prokaryotic initiation factors, reflecting the greater biological complexity of eukaryotic translation. There are at least twelve eukaryotic initiation factors, composed of many more polypeptides, and these are described below.

<span class="mw-page-title-main">Eukaryotic translation termination factor 1</span> Protein-coding gene in the species Homo sapiens

Eukaryotic translation termination factor 1 (eRF1), also known as TB3-1, is a protein that in humans is encoded by the ETF1 gene.

Elongation factor 4 (EF-4) is an elongation factor that is thought to back-translocate on the ribosome during the translation of RNA to proteins. It is found near-universally in bacteria and in eukaryotic endosymbiotic organelles including the mitochondria and the plastid. Responsible for proofreading during protein synthesis, EF-4 is a recent addition to the nomenclature of bacterial elongation factors.

Protein-synthesizing GTPases are enzymes involved in mRNA translation into protein by the ribosome, with systematic name GTP phosphohydrolase (mRNA-translation-assisting). They usually include translation initiation factors such as IF-2 and translation elongation factors such as EF-Tu.

<span class="mw-page-title-main">Prokaryotic large ribosomal subunit</span>

50S is the larger subunit of the 70S ribosome of prokaryotes, i.e. bacteria and archaea. It is the site of inhibition for antibiotics such as macrolides, chloramphenicol, clindamycin, and the pleuromutilins. It includes the 5S ribosomal RNA and 23S ribosomal RNA.

<span class="mw-page-title-main">EF-G</span> Prokaryotic elongation factor

EF-G is a prokaryotic elongation factor involved in protein translation. As a GTPase, EF-G catalyzes the movement (translocation) of transfer RNA (tRNA) and messenger RNA (mRNA) through the ribosome.

<span class="mw-page-title-main">GSPT2</span> Protein-coding gene in the species Homo sapiens

Eukaryotic peptide chain release factor GTP-binding subunit ERF3B is an enzyme that in humans is encoded by the GSPT2 gene.

<span class="mw-page-title-main">Protein synthesis inhibitor</span> Inhibitors of translation

A protein synthesis inhibitor is a compound that stops or slows the growth or proliferation of cells by disrupting the processes that lead directly to the generation of new proteins.

Translational regulation refers to the control of the levels of protein synthesized from its mRNA. This regulation is vastly important to the cellular response to stressors, growth cues, and differentiation. In comparison to transcriptional regulation, it results in much more immediate cellular adjustment through direct regulation of protein concentration. The corresponding mechanisms are primarily targeted on the control of ribosome recruitment on the initiation codon, but can also involve modulation of peptide elongation, termination of protein synthesis, or ribosome biogenesis. While these general concepts are widely conserved, some of the finer details in this sort of regulation have been proven to differ between prokaryotic and eukaryotic organisms.

Sup45p is the Saccharomyces cerevisiae eukaryotic translation termination factor. More specifically, it is the yeast eukaryotic release factor 1 (eRF1). Its job is to recognize stop codons in RNA and bind to them. It binds to the Sup35p protein and then takes on the shape of a tRNA molecule so that it can safety incorporate itself into the A site of the Ribosome to disruptits flow and "release" the protein and end translation.

The P-site is the second binding site for tRNA in the ribosome. The other two sites are the A-site (aminoacyl), which is the first binding site in the ribosome, and the E-site (exit), the third. During protein translation, the P-site holds the tRNA which is linked to the growing polypeptide chain. When a stop codon is reached, the peptidyl-tRNA bond of the tRNA located in the P-site is cleaved releasing the newly synthesized protein. During the translocation step of the elongation phase, the mRNA is advanced by one codon, coupled to movement of the tRNAs from the ribosomal A to P and P to E sites, catalyzed by elongation factor EF-G.

References

  1. ↑ Capecchi MR (September 1967). "Polypeptide chain termination in vitro: isolation of a release factor". Proceedings of the National Academy of Sciences of the United States of America. 58 (3): 1144–1151. Bibcode:1967PNAS...58.1144C. doi: 10.1073/pnas.58.3.1144 . PMC   335760 . PMID   5233840.
  2. ↑ Scolnick E, Tompkins R, Caskey T, Nirenberg M (October 1968). "Release factors differing in specificity for terminator codons". Proceedings of the National Academy of Sciences of the United States of America. 61 (2): 768–774. Bibcode:1968PNAS...61..768S. doi: 10.1073/pnas.61.2.768 . PMC   225226 . PMID   4879404.
  3. ↑ Brown CM, Tate WP (December 1994). "Direct recognition of mRNA stop signals by Escherichia coli polypeptide chain release factor two". The Journal of Biological Chemistry. 269 (52): 33164–33170. doi: 10.1016/S0021-9258(20)30112-5 . PMID   7806547.
  4. ↑ Scarlett DJ, McCaughan KK, Wilson DN, Tate WP (April 2003). "Mapping functionally important motifs SPF and GGQ of the decoding release factor RF2 to the Escherichia coli ribosome by hydroxyl radical footprinting. Implications for macromolecular mimicry and structural changes in RF2". The Journal of Biological Chemistry. 278 (17): 15095–15104. doi: 10.1074/jbc.M211024200 . PMID   12458201.
  5. ↑ Jakobsen CG, Segaard TM, Jean-Jean O, Frolova L, Justesen J (2001). "[Identification of a novel termination release factor eRF3b expressing the eRF3 activity in vitro and in vivo]". Molekuliarnaia Biologiia. 35 (4): 672–681. PMID   11524954.
  6. 1 2 Weaver RF (2005). Molecular Biology . New York, NY: McGraw-Hill. pp.  616–621. ISBN   978-0-07-284611-9.
  7. ↑ Saito K, Kobayashi K, Wada M, Kikuno I, Takusagawa A, Mochizuki M, et al. (November 2010). "Omnipotent role of archaeal elongation factor 1 alpha (EF1α in translational elongation and termination, and quality control of protein synthesis". Proceedings of the National Academy of Sciences of the United States of America. 107 (45): 19242–19247. Bibcode:2010PNAS..10719242S. doi: 10.1073/pnas.1009599107 . PMC   2984191 . PMID   20974926.
  8. ↑ Buckingham RH, Grentzmann G, Kisselev L (May 1997). "Polypeptide chain release factors". Molecular Microbiology. 24 (3): 449–456. doi: 10.1046/j.1365-2958.1997.3711734.x . PMID   9179839. Standard methods of sequence comparison do not show significant similarity between the prokaryotic factors RF1/2 and RF1
  9. ↑ Kisselev L (January 2002). "Polypeptide Release Factors in Prokaryotes and Eukaryotes". Structure. 10 (1): 8–9. doi: 10.1016/S0969-2126(01)00703-1 . PMID   11796105.
  10. ↑ Inagaki Y, Ford Doolittle W (June 2000). "Evolution of the eukaryotic translation termination system: origins of release factors". Molecular Biology and Evolution. 17 (6): 882–889. doi:10.1093/oxfordjournals.molbev.a026368. PMID   10833194.
  11. ↑ Duarte I, Nabuurs SB, Magno R, Huynen M (November 2012). "Evolution and diversification of the organellar release factor family". Molecular Biology and Evolution. 29 (11): 3497–3512. doi:10.1093/molbev/mss157. PMC   3472500 . PMID   22688947.
  12. 1 2 3 4 Zhou J, Korostelev A, Lancaster L, Noller HF (December 2012). "Crystal structures of 70S ribosomes bound to release factors RF1, RF2 and RF3". Current Opinion in Structural Biology. 22 (6): 733–742. doi:10.1016/j.sbi.2012.08.004. PMC   3982307 . PMID   22999888.
  13. 1 2 Taylor D, Unbehaun A, Li W, Das S, Lei J, Liao HY, et al. (November 2012). "Cryo-EM structure of the mammalian eukaryotic release factor eRF1-eRF3-associated termination complex". Proceedings of the National Academy of Sciences of the United States of America. 109 (45): 18413–18418. Bibcode:2012PNAS..10918413T. doi: 10.1073/pnas.1216730109 . PMC   3494903 . PMID   23091004.
  14. 1 2 3 des Georges A, Hashem Y, Unbehaun A, Grassucci RA, Taylor D, Hellen CU, et al. (March 2014). "Structure of the mammalian ribosomal pre-termination complex associated with eRF1.eRF3.GDPNP". Nucleic Acids Research. 42 (5): 3409–3418. doi:10.1093/nar/gkt1279. PMC   3950680 . PMID   24335085.
  15. ↑ Fu Z, Indrisiunaite G, Kaledhonkar S, Shah B, Sun M, Chen B, et al. (June 2019). "The structural basis for release-factor activation during translation termination revealed by time-resolved cryogenic electron microscopy". Nature Communications. 10 (1): 2579. Bibcode:2019NatCo..10.2579F. doi:10.1038/s41467-019-10608-z. PMC   6561943 . PMID   31189921.
  16. ↑ Indrisiunaite G, Pavlov MY, Heurgué-Hamard V, Ehrenberg M (May 2015). "On the pH dependence of class-1 RF-dependent termination of mRNA translation". Journal of Molecular Biology. Translation: Regulation and Dynamics. 427 (9): 1848–1860. doi: 10.1016/j.jmb.2015.01.007 . PMID   25619162.
  17. ↑ Pavlov MY, Antoun A, Lovmar M, Ehrenberg M (June 2008). "Complementary roles of initiation factor 1 and ribosome recycling factor in 70S ribosome splitting". The EMBO Journal. 27 (12): 1706–1717. doi:10.1038/emboj.2008.99. PMC   2435134 . PMID   18497739.
  18. ↑ Kobayashi K, Saito K, Ishitani R, Ito K, Nureki O (October 2012). "Structural basis for translation termination by archaeal RF1 and GTP-bound EF1α complex". Nucleic Acids Research. 40 (18): 9319–9328. doi:10.1093/nar/gks660. PMC   3467058 . PMID   22772989.
  19. ↑ Becker T, Franckenberg S, Wickles S, Shoemaker CJ, Anger AM, Armache JP, et al. (February 2012). "Structural basis of highly conserved ribosome recycling in eukaryotes and archaea". Nature. 482 (7386): 501–506. Bibcode:2012Natur.482..501B. doi:10.1038/nature10829. PMC   6878762 . PMID   22358840.
  20. ↑ Hellen CU (October 2018). "Translation Termination and Ribosome Recycling in Eukaryotes". Cold Spring Harbor Perspectives in Biology. 10 (10): a032656. doi:10.1101/cshperspect.a032656. PMC   6169810 . PMID   29735640.
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