Ribosomes terminate translation when release factors (RFs) recognize an mRNA stop codon in the A (aminoacyl-tRNA) site (Brenner et al., 1967; Brenner et al., 1965). Upon stop-codon recognition, the release factor hydrolyzes peptidyl-tRNA in the P site of the large subunit, thus releasing nascent protein from the ribosome (Capecchi, 1967a; Capecchi, 1967b; Scolnick et al., 1968; Tompkins et al., 1970). In bacteria, RF1 recognizes UAA or UAG, and RF2 recognizes UAA or UGA codons (Nakamura et al., 1995). Following peptide release, RF1 or RF2 must dissociate from the ribosome to allow the ribosome and release factor to recycle. While structural studies have provided snapshots of some peptidyl-tRNA hydrolysis steps (reviewed in Dunkle and Cate, 2010; Korostelev, 2011; Ramakrishnan, 2011; Rodnina, 2018; Youngman et al., 2008), recent biophysical and biochemical findings suggest a highly dynamic series of termination events (Adio et al., 2018; Indrisiunaite et al., 2015; Prabhakar et al., 2017; Sternberg et al., 2009), which have not been structurally visualized.

In the pre-termination (pre-hydrolysis) state, the substrate peptidyl-tRNA resides in a non-rotated (classical) conformation of 70S ribosome, with its anticodon stem loop bound to the mRNA codon in the P site of the small 30S subunit and the 3′-terminal CCA-peptidyl moiety in the P site of the large 50S subunit (Agrawal et al., 2000; Ermolenko et al., 2007). Cryogenic electron microscopy (cryo-EM) and X-ray studies reveal RF1 or RF2 bound in the A site next to P tRNA (Klaholz et al., 2003; Petry et al., 2005; Rawat et al., 2006; Rawat et al., 2003), and provide insight into pre-hydrolysis (Jin et al., 2010) and post-hydrolysis states (Korostelev et al., 2008; Korostelev et al., 2010; Laurberg et al., 2008; Weixlbaumer et al., 2008). The release factor’s codon-recognition domain (domain 2) binds the stop codon in the 30S decoding center. The catalytic GGQ motif (glycine-glycine-glutamine of domain 3) inserts into the 50S peptidyl-transferase center (PTC) adjacent to the CCA end of P-tRNA. Ribosomes retain the non-rotated conformation in RF-bound structures, including those with mutated or post-translationally modified RFs (Pierson et al., 2016; Santos et al., 2013; Zeng and Jin, 2018) or when RFs are perturbed by the termination inhibitor blasticidin S (Svidritskiy and Korostelev, 2018a; Svidritskiy and Korostelev, 2018b) or by a sense codon in the A site (Svidritskiy et al., 2018). The catalytic GGQ loop adopts the same compact conformation comprising a short α-helix in pre-hydrolysis-like and post-hydrolysis structures, as discussed below.

Ribosome and RF dynamics are implicated in post-hydrolysis steps. In E. coli, RF1 dissociation is assisted by the non-essential GTPase RF3 (Freistroffer et al., 1997; Koutmou et al., 2014), whereas RF2 can spontaneously dissociate from the ribosome (Adio et al., 2018). RF3 induces rotation of the small subunit relative to the large subunit by ~9 degrees in the absence of RF1 and RF2 (Ermolenko et al., 2007; Zhou et al., 2012) and stabilizes a ‘hybrid’ state of deacylated tRNA (P/E) (Gao et al., 2007; Jin et al., 2011). In the P/E conformation, the anticodon stem loop remains in the 30SS P site, while the elbow and acceptor arm bind the L1 stalk and E (exit) site of the 50S subunit. A recent cryo-EM study (Graf et al., 2018) showed that if RF1 is locked on the post-hydrolysis ribosome by antibacterial peptide apidaecin 137 (Florin et al., 2017), the ribosome can undergo intersubunit rotation in the presence of both RF1 and RF3. By contrast, biochemical experiments showed that locking RF1 on the ribosome by inactivating its catalytic motif (GGQ->GAQ) prevented RF1 recycling by RF3 from the pre-hydrolysis ribosome (Koutmou et al., 2014), echoing the deficiency of mutant RF2 (GGQ->GAQ) in stimulating GTP hydrolysis by RF3 (Zavialov et al., 2002). These and other (Adio et al., 2018) observations indicate that peptidyl-tRNA hydrolysis is critical for RF dissociation, likely due to propensity of deacylated tRNA for exiting the P site and sampling the P/E hybrid state (Moazed and Noller, 1989) via spontaneous intersubunit rotation (Cornish et al., 2008). Nevertheless, RF3 is dispensable for growth of Escherichia coli (Grentzmann et al., 1994; Mikuni et al., 1994), and its expression is not conserved in bacteria (Leipe et al., 2002). For example, RF3 is not present in the thermophilic model organisms of the Thermus and Thermatoga genera and in infectious Chlamydiales and Spirochaetae. This means that both RF1 and RF2 are capable of performing a complete round of termination independently of RF3. Indeed, recent Förster Resonance Energy Transfer (FRET) studies demonstrate spontaneous RF1 and RF2 dissociation after peptidyl-tRNA hydrolysis (Prabhakar et al., 2017), coupled with transient RF sampling of the rotated ribosomes (Adio et al., 2018). The structural basis for RF dissociation, however, remains incompletely understood because RFs have not been structurally visualized on ribosomes with rearranged/rotated subunits in the absence of RF3 and inhibitors.

In this work, we used ensemble cryo-EM, as described earlier (Abeyrathne et al., 2016; Loveland et al., 2017), employing 3D maximum likelihood classification/sorting of particle images (Lyumkis et al., 2013; Scheres, 2010; Scheres et al., 2007) to visualize the structural dynamics of RF2 on the 70S ribosome (Figure 1, Figure 1—figure supplement 1, Figure 1—figure supplement 2 and Figure 1—figure supplement 3). The structures capture RF2 and tRNA in distinct 70S conformational states (Table 1). They reveal an unexpected rearrangement of the catalytic GGQ loop, which suggests that RF2 helps direct nascent protein out of the ribosome. Our findings reconcile biophysical observations and, together with other structural studies, allow a reconstruction of the termination course by RF2, from initial binding to dissociation.

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We formed a termination complex using E. coli 70S ribosomes and RF2 to catalyze the hydrolysis of substrate N-formyl-methionyl-tRNA^fMet (fMet-tRNA^fMet) bound to mRNA carrying an AUG codon in the P site and a UGA stop codon in the A site. Although the stop codon is placed immediately after the start codon, unlike in cellular mRNAs, such model termination complexes have been successfully used in numerous structural, biochemical and biophysical studies (Adio et al., 2018; Casy et al., 2018; Koutmou et al., 2014; Kuhlenkoetter et al., 2011; Laurberg et al., 2008; Pallesen et al., 2013; Pierson et al., 2016; Shi and Joseph, 2016; Sternberg et al., 2009). Indeed, under the conditions used for cryo-EM sample preparation in this work, RF2 supports hydrolysis of fMet-tRNA^fMet and release of fMet (Figure 1—figure supplement 4), demonstrating activity of the ribosome and RF2 in our sample. Maximum-likelihood classification of cryo-EM data in FREALIGN (Lyumkis et al., 2013) yielded five structures at average resolutions of 3.3 Å to 4.4 Å (Table 2; Figure 1). The structures differ by the degrees of intersubunit rotation and 30S head rotation (swivel), and are consistent with the following termination stages: pre-termination (i.e., no RF2; Structure I); peptidyl-tRNA hydrolysis by RF2 (Structure II); tRNA exit from the PTC (Structure III); destabilization of the catalytic domain of RF2 (Structure IV); and post-termination (i.e., after RF2 dissociates; Structure V).

Pre-termination-like Structure I contains tRNA in the P site with its 3′-CCA end positioned in the peptidyl transferase center (Figure 1A). Although the tRNA is likely partially or completely deacylated due to the action of RF2, the tRNA and ribosome conformations closely resemble those in pre-termination aminoacyl- and peptidyl-tRNA complexes (Adio et al., 2018; Cornish et al., 2008; Jin et al., 2010; Polikanov et al., 2014). The 30S subunit adopts a conformation similar to crystal structures of non-rotated ribosomes with or without RFs (Jin et al., 2010; Korostelev et al., 2008; Korostelev et al., 2006; Korostelev et al., 2010; Laurberg et al., 2008; Selmer et al., 2006; Weixlbaumer et al., 2008), and exhibits a slight ~1.5° rotation relative to the 70S•RF2 crystal structure formed with the UGA stop codon (Weixlbaumer et al., 2008).

RF2 is bound in Structures II to IV (Figure 1B–D). The 70S intersubunit rotation ranges from ~1.5° in Structure II to ~7.5° in Structure IV (Table 1). This nearly full range of rotation is typical of translocation complexes with tRNA (Agirrezabala et al., 2008; Agirrezabala et al., 2012; Dunkle et al., 2011; Fischer et al., 2010; Julián et al., 2008) and elongation factor G (EF-G) (Brilot et al., 2013; Chen et al., 2013; Frank and Agrawal, 2000; Gao et al., 2009; Pulk and Cate, 2013; Tourigny et al., 2013; Zhou et al., 2013).

The most well-resolved 3.3 Å resolution Structure II (Figure 1B) accounts for the largest population of RF2-bound ribosome particles in the sample (Table 2), consistent with stabilization of the non-rotated 30S states by RF2 observed in FRET studies (Casy et al., 2018; Prabhakar et al., 2017; Sternberg et al., 2009). The overall extended conformation of RF2 resembles that seen in published 70S•RF2 structures (Korostelev et al., 2008; Weixlbaumer et al., 2008) with its codon-recognition superdomain (domains 2 and 4; Figure 1B) bound to the UGA stop codon in the decoding center of the 30S subunit and its catalytic domain 3 inserted into the peptidyl-transferase center.

Structure II features RF2 with a dramatically rearranged catalytic loop, coinciding with the lack of density for the CCA end of the P tRNA. In previous pre- and post-hydrolysis-like structures, the P-tRNA CCA end binds the PTC, and RF2 forms a compact catalytic loop (residues 245–258). In these previous structures, the catalytic ²⁵⁰GGQ²⁵² motif resides at the tip of a short α-helix, adjacent to the terminal tRNA nucleotide A76 (Figure 2A, light shades; (Jin et al., 2010; Korostelev et al., 2008; Laurberg et al., 2008; Weixlbaumer et al., 2008). The α-helical region is held in place by A2602 of 23S ribosomal RNA (rRNA), which is critical for termination efficiency (Amort et al., 2007; Polacek et al., 2003). In Structure II, however, the catalytic loop adopts an extended β-hairpin conformation that reaches ~10 Å deeper into the peptide tunnel (Figure 2A–B). To accommodate the extended β-hairpin, nucleotides G2505 and U2506 of 23S rRNA are shifted to widen the PTC (Figure 2—figure supplement 1A). The center of the β-hairpin occupies the position normally held by A76 of P tRNA (Figure 2A). The lack of density for the CCA moiety thus suggests that the 3′ end of P tRNA is at least partially released from the PTC. Indeed, formation of the β-hairpin and release of the CCA moiety is accompanied by a slight shift in the P-tRNA acceptor arm away from the PTC and by ~160° rotation of nucleotide A2602 to form a Hoogsteen base pair with C1965 at helix 71 of 23S rRNA (Figure 2A and Figure 2Figure 2—figure supplement 1B). The cryo-EM map shows strong density for the extended β strands (Figure 2C), but weaker density for the GGQ residues at the tip of the hairpin, consistent with conformational heterogeneity of the flexible glycine backbone (Figure 2—figure supplement 2). Nevertheless, the tip of the β-hairpin plugs the narrowest region of the peptide tunnel at A2062 of 23S rRNA (Figure 2B). Thus, Structure II appears to represent a previously unseen post-peptide-release state with deacyl-tRNA dissociating from the PTC.

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In Structure III (Figure 1C), a 5° rotation of the 30S subunit shifts the tRNA elbow by ~10 Å along protein L5 toward the E site. As a result, the P-tRNA acceptor arm pulls further out of the 50S P site, and the end of the acceptor arm helix points at helix α7 of RF2 near positively charged Lys 282 and Lys 289. The CCA moiety remains unresolved. The 5° intersubunit rotation also causes a slight 2° rotation of RF2 domain 3 relative to domain 2, but the overall conformation of RF2 remains similar to that in Structure II, with the tip of the extended β-hairpin plugging the peptide exit tunnel.

In Structure IV, the 30S subunit is rotated 7.5° and the P-tRNA elbow is bound to the L1 stalk, an interaction normally observed with the P/E or E tRNA (Dunkle et al., 2011; Korostelev et al., 2006; Selmer et al., 2006). Unlike P/E tRNA, however, the acceptor arm is oriented between the 50S P and E sites (Figure 1D), suggesting that the tRNA adopts a transient state on its way toward the hybrid P/E conformation. This transition coincides with an ~3° rotation (swivel) of the 30S head, characteristic of intermediate stages of tRNA and mRNA translocation (Abeyrathne et al., 2016; Ratje et al., 2010). The codon-recognition superdomain of RF2 in its canonical conformation moves with the 30S subunit, causing the catalytic domain to shift by ~5 Å from its position in Structure II (Figure 3A). Indeed, reduced density for the catalytic domain and domain 1, which bridges the 30S and 50S subunits at the A site periphery (Figure 1B), suggests that intersubunit rotation has destabilized RF2. In Structure IV, therefore, RF2 appears to be poised for dissociation and collapsing toward the compact conformation adopted by free RFs (Figure 3A) (JCSG, 2005; Shin et al., 2004; Vestergaard et al., 2001; Zoldák et al., 2007).

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Structure V is incompatible with RF2 binding and provides further insights into RF2 dissociation. The rotated 70S•tRNA structure resembles hybrid P/E states, with the acceptor arm of tRNA directed toward the E site of 50S (Figure 1E). However, the central intersubunit bridge B2a—formed by helix 69 (H69, nt. 1906–1924) of 23S rRNA—is disassembled (Figure 3B–C). In RF-bound structures (including Structures I to IV, above), the tip of H69 (at nt 1913) locks into the decoding center in a termination-specific arrangement (Korostelev et al., 2008; Korostelev et al., 2010; Laurberg et al., 2008; Weixlbaumer et al., 2008). H69 interacts with the switch loop of RFs, where the codon-recognition superdomain connects to catalytic domain 3 (e.g., Structure II, Figure 3B). This interaction directs the catalytic domain toward the PTC (Korostelev et al., 2010; Laurberg et al., 2008) and thus defines the efficiency and accuracy of release factors (Svidritskiy and Korostelev, 2018a). In Structure V, however, H69 is disengaged from the decoding center and moved ~15 Å toward the large subunit to pack against H71 near A2602 (nt 1945–1961; Figure 3C). The new position of H69 would clash with extended or compact conformations of RF2 (Figure 3—figure supplement 1), if the release factor had remained bound to the decoding center.

Release of RF2 also appears to be coupled to rotation (swivel) of the 30S head domain. In RF2-bound structures, the conserved codon-recognition ²⁰⁵SPF²⁰⁷ motif (serine-proline-phenylalanine of domain 2) of RF2 interacts with the stop codon and the 30S head near bulged nucleotide C1054 of 16S rRNA. In Structure V, a 6° rotation of the 30S head shifts the position of C1054 by 5 Å from its position in RF2-bound Structure IV, consistent with disassembly of RF2 contacts. The disruption of decoding-center interactions due to repositioning of H69 and head swivel therefore explain the absence of RF2 in Structure V.

Mechanism of translation termination by RF2

Our termination structures allow us to reconstruct a dynamic pathway for termination (Figure 4 and Animation 1) and reconcile biophysical and biochemical findings (Adio et al., 2018; Casy et al., 2018; Prabhakar et al., 2017; Sternberg et al., 2009). Recent structural studies revealed compact RF conformations on non-rotated 70S ribosomes (Fu et al., 2018; Svidritskiy and Korostelev, 2018a) that suggest how RFs open at early stages of codon recognition (He and Green, 2010; Hetrick et al., 2009; Trappl and Joseph, 2016). The short-lived transition from compact to open conformation(s) lasts tens of milliseconds (Fu et al., 2018), and compact RF2 conformations were not captured in our sample. Compact RF2 was also visualized by cryo-EM on the 70S ribosome in the presence of truncated mRNA and alternative rescue factor A (ArfA) (Demo et al., 2017; James et al., 2016), suggesting a conserved pathway for initial binding of RFs to non-rotated ribosomes. Together with these recent studies, our work provides an expanded structural view of the termination reaction – from initial codon recognition to peptide release to RF2 dissociation (Figure 4).

Figure 4

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Termination begins when release factor recognizes a stop codon in the A site of a non-rotated (or slightly rotated) 70S ribosome carrying peptidyl-tRNA (Figure 4A). A compact release factor recognizes the stop codon, while its catalytic domain is kept 60 to 70 Å away from the PTC (Fu et al., 2018). Separation of the codon-recognition and catalytic functions during initial binding of RF underlies high accuracy of termination (He and Green, 2010; Laurberg et al., 2008; Svidritskiy and Korostelev, 2018a; Trappl and Joseph, 2016). Upon binding the decoding center, steric hindrance with P-tRNA forces the RF catalytic domain to undock from domain two and sample the intersubnit space (Figure 4B) (discussed in Svidritskiy and Korostelev, 2018a). The ribosome remains non-rotated because peptidyl-tRNA bridges the P sites of 30S and 50S subunits. Interactions between the switch loop of RF and the decoding center and H69 direct the catalytic domain into the PTC. The GGQ catalytic loop is placed next to the scissile ester bond of peptidyl-tRNA and stabilized by A2602 (Jin et al., 2010; Laurberg et al., 2008), so that the Gln backbone amide of the GGQ motif can catalyze peptidyl-tRNA hydrolysis (Figure 4C) (Korostelev et al., 2008; Santos et al., 2013). These ribosome and RF2 rearrangements complete the first stage of termination leading to peptidyl-tRNA hydrolysis—an irreversible step that separates the newly made protein from P-site tRNA and prevents further elongation.

To be recycled, the ribosome must release the nascent protein and RF. The deacylated CCA end of the tRNA exits the 50S P site, which allows movement of A2602. No longer stabilized by the tRNA and A2602, the GGQ loop reorganizes into an extended β-hairpin that blocks the peptide tunnel (Structure II), thus biasing the diffusion of nascent peptide toward the exit of the peptide tunnel at the surface of the 50S subunit (Figure 4D). Furthermore, the exit of the CCA end from the P site supports formation of the P/E-tRNA hybrid conformation via spontaneous intersubunit rotation (Structures III and IV), which is also essential for translocation of tRNA-mRNA (reviewed in Ling and Ermolenko, 2016; Noller et al., 2017). The large counter-clockwise rotation of the small subunit destabilizes the catalytic domain of RF2 (Structure IV), so that it begins to collapse toward domain 2 bound to the 30S subunit (Figure 4E). Rotation of the 30S head and detachment of H69 from the decoding center force dissociation of RF2 from the A site of the 30S subunit (Structure V, Figure 4F).

Remarkably similar extents of intersubunit rotation and head rotation were observed in recent RF1-bound 70S structures formed with RF3•GDPNP in the presence of apidaecin (which stalls RF1 on the ribosome). In the presence of stalled RF1, however, the intersubunit bridge remains attached to the decoding center (Graf et al., 2018). Thus, the ribosome samples globally similar rotation states with RF1 and RF2, with or without RF3. RF3 stabilizes the rotated state (Ermolenko et al., 2007; Gao et al., 2007; Jin et al., 2011; Zhou et al., 2012) to promote RF release. However, the role of RF3 in termination is dispensable because intersubunit rotation is an inherent— that is spontaneous and thermally driven—property of the ribosome (Cornish et al., 2008). Cold sensitivity of RF3-deletion strains (Nichols et al., 2011; O'Connor, 2015) perhaps manifests the dependence on RF3 due to perturbation of the intersubunit dynamics at lower temperatures. In the fully rotated state, the post-termination ribosome becomes a substrate for subunit dissociation by EF-G and recycling factor RRF (Agrawal et al., 2004; Dunkle et al., 2011; Fu et al., 2016; Prabhakar et al., 2017). Structure V is consistent with structural studies showing ribosome recycling factor (RRF) bound to ribosomes with a dislodged intersubunit bridge B2a (Borovinskaya et al., 2007; Pai et al., 2008), which helps split the ribosome into subunits. Translation termination and recycling of the release factors and the ribosome therefore rely on the spontaneous ribosome dynamics (Adio et al., 2018; Cornish et al., 2008; Ermolenko et al., 2007; Prabhakar et al., 2017; Sternberg et al., 2009), coupled with local rearrangements of the universally conserved elements of the peptidyl-transferase and decoding centers.

Structural models have been deposited in PDB under the accession codes 6OFX, 6OG7, 6OGF, 6OGG, 6OGI. Cryo-EM data have been deposited to EMDB under the accession codes EMD-20048, EMD-20052, EMD-20056, EMD-20057, EMD-20058.

1. 1. Svidritskiy E
   2. Demo G
   3. Loveland AB
   4. Xu C
   5. Korostelev AA

   (2019) Protein Data Bank

   ID 6OFX. Non-rotated ribosome (Structure I).

   http://www.rcsb.org/structure/6OFX
2. 1. Svidritskiy E
   2. Demo G
   3. Loveland AB
   4. Xu C
   5. Korostelev AA

   (2019) Electron Microscopy Data Bank

   ID EMD-20048. Non-rotated ribosome (Structure I).

   https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-20048
3. 1. Svidritskiy E
   2. Demo G
   3. Loveland AB
   4. Xu C
   5. Korostelev AA

   (2019) Protein Data Bank

   ID 6OG7. 70S termination complex with RF2 bound to the UGA codon. Non-rotated ribosome with RF2 bound (Structure II).

   http://www.rcsb.org/structure/6OG7
4. 1. Svidritskiy E
   2. Demo G
   3. Loveland AB
   4. Xu C
   5. Korostelev AA

   (2019) Protein Data Bank

   ID 6OGF. 70S termination complex with RF2 bound to the UGA codon. Partially rotated ribosome with RF2 bound (Structure III).

   http://www.rcsb.org/structure/6OGF
5. 1. Svidritskiy E
   2. Demo G
   3. Loveland AB
   4. Xu C
   5. Korostelev AA

   (2019) Protein Data Bank

   ID 6OGI. 70S termination complex with the UGA stop codon. Rotated ribosome conformation (Structure V).

   http://www.rcsb.org/structure/6OGI
6. 1. Svidritskiy E
   2. Demo G
   3. Loveland AB
   4. Xu C
   5. Korostelev AA

   (2019) Protein Data Bank

   ID 6OGG. 70S termination complex with RF2 bound to the UGA codon. Rotated ribosome with RF2 bound (Structure IV).

   http://www.rcsb.org/structure/6OGG
7. 1. Svidritskiy E
   2. Demo G
   3. Loveland AB
   4. Xu C
   5. Korostelev AA

   (2019) Electron Microscopy Data Bank

   ID EMD-20052. 70S termination complex with RF2 bound to the UGA codon. Non-rotated ribosome with RF2 bound (Structure II).

   https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-20052
8. 1. Svidritskiy E
   2. Demo G
   3. Loveland AB
   4. Xu C
   5. Korostelev AA

   (2019) Electron Microscopy Data Bank

   ID EMD-20056. 70S termination complex with RF2 bound to the UGA codon. Partially rotated ribosome with RF2 bound (Structure III).

   https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-20056
9. 1. Svidritskiy E
   2. Demo G
   3. Loveland AB
   4. Xu C
   5. Korostelev AA

   (2019) Electron Microscopy Data Bank

   ID EMD-20057. 70S termination complex with RF2 bound to the UGA codon. Rotated ribosome with RF2 bound (Structure IV).

   https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-20057
10. 1. Svidritskiy E
    2. Demo G
    3. Loveland AB
    4. Xu C
    5. Korostelev AA

    (2019) Electron Microscopy Data Bank

    ID EMD-20058. 70S termination complex with the UGA stop codon. Rotated ribosome conformation (Structure V).

    https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-20058

1. 1. Abeyrathne PD
   2. Koh CS
   3. Grant T
   4. Grigorieff N
   5. Korostelev AA

   (2016) Ensemble cryo-EM uncovers inchworm-like translocation of a viral IRES through the ribosome

   eLife 5:e14874.

   https://doi.org/10.7554/eLife.14874

   • PubMed
   • Google Scholar
2. 1. Adams PD
   2. Grosse-Kunstleve RW
   3. Hung LW
   4. Ioerger TR
   5. McCoy AJ
   6. Moriarty NW
   7. Read RJ
   8. Sacchettini JC
   9. Sauter NK
   10. Terwilliger TC

   (2002) PHENIX: building new software for automated crystallographic structure determination

   Acta Crystallographica Section D Biological Crystallography 58:1948–1954.

   https://doi.org/10.1107/S0907444902016657

   • PubMed
   • Google Scholar
3. 1. Adams PD
   2. Afonine PV
   3. Bunkóczi G
   4. Chen VB
   5. Davis IW
   6. Echols N
   7. Headd JJ
   8. Hung LW
   9. Kapral GJ
   10. Grosse-Kunstleve RW
   11. McCoy AJ
   12. Moriarty NW
   13. Oeffner R
   14. Read RJ
   15. Richardson DC
   16. Richardson JS
   17. Terwilliger TC
   18. Zwart PH

   (2010) PHENIX: a comprehensive Python-based system for macromolecular structure solution

   Acta Crystallographica Section D Biological Crystallography 66:213–221.

   https://doi.org/10.1107/S0907444909052925

   • PubMed
   • Google Scholar
4. 1. Adio S
   2. Sharma H
   3. Senyushkina T
   4. Karki P
   5. Maracci C
   6. Wohlgemuth I
   7. Holtkamp W
   8. Peske F
   9. Rodnina MV

   (2018) Dynamics of ribosomes and release factors during translation termination in E. coli

   eLife 7:e34252.

   https://doi.org/10.7554/eLife.34252

   • PubMed
   • Google Scholar
5. 1. Agirrezabala X
   2. Lei J
   3. Brunelle JL
   4. Ortiz-Meoz RF
   5. Green R
   6. Frank J

   (2008) Visualization of the hybrid state of tRNA binding promoted by spontaneous ratcheting of the ribosome

   Molecular Cell 32:190–197.

   https://doi.org/10.1016/j.molcel.2008.10.001

   • PubMed
   • Google Scholar
6. 1. Agirrezabala X
   2. Liao HY
   3. Schreiner E
   4. Fu J
   5. Ortiz-Meoz RF
   6. Schulten K
   7. Green R
   8. Frank J

   (2012) Structural characterization of mRNA-tRNA translocation intermediates

   PNAS 109:6094–6099.

   https://doi.org/10.1073/pnas.1201288109

   • PubMed
   • Google Scholar
7. 1. Agrawal RK
   2. Spahn CM
   3. Penczek P
   4. Grassucci RA
   5. Nierhaus KH
   6. Frank J

   (2000) Visualization of tRNA movements on the Escherichia coli 70S ribosome during the elongation cycle

   The Journal of Cell Biology 150:447–460.

   https://doi.org/10.1083/jcb.150.3.447

   • PubMed
   • Google Scholar
8. 1. Agrawal RK
   2. Sharma MR
   3. Kiel MC
   4. Hirokawa G
   5. Booth TM
   6. Spahn CM
   7. Grassucci RA
   8. Kaji A
   9. Frank J

   (2004) Visualization of ribosome-recycling factor on the Escherichia coli 70S ribosome: functional implications

   PNAS 101:8900–8905.

   https://doi.org/10.1073/pnas.0401904101

   • PubMed
   • Google Scholar
9. 1. Amort M
   2. Wotzel B
   3. Bakowska-Zywicka K
   4. Erlacher MD
   5. Micura R
   6. Polacek N

   (2007) An intact ribose moiety at A2602 of 23S rRNA is key to trigger peptidyl-tRNA hydrolysis during translation termination

   Nucleic Acids Research 35:5130–5140.

   https://doi.org/10.1093/nar/gkm539

   • PubMed
   • Google Scholar
10. 1. Borovinskaya MA
    2. Pai RD
    3. Zhang W
    4. Schuwirth BS
    5. Holton JM
    6. Hirokawa G
    7. Kaji H
    8. Kaji A
    9. Cate JH

    (2007) Structural basis for aminoglycoside inhibition of bacterial ribosome recycling

    Nature Structural & Molecular Biology 14:727–732.

    https://doi.org/10.1038/nsmb1271

    • PubMed
    • Google Scholar
11. 1. Brenner S
    2. Stretton AO
    3. Kaplan S

    (1965) Genetic code: the 'nonsense' triplets for chain termination and their suppression

    Nature 206:994–998.

    https://doi.org/10.1038/206994a0

    • PubMed
    • Google Scholar
12. 1. Brenner S
    2. Barnett L
    3. Katz ER
    4. Crick FH

    (1967) UGA: a third nonsense triplet in the genetic code

    Nature 213:449–450.

    https://doi.org/10.1038/213449a0

    • PubMed
    • Google Scholar
13. 1. Brilot AF
    2. Korostelev AA
    3. Ermolenko DN
    4. Grigorieff N

    (2013) Structure of the ribosome with elongation factor G trapped in the pretranslocation state

    PNAS 110:20994–20999.

    https://doi.org/10.1073/pnas.1311423110

    • PubMed
    • Google Scholar
14. 1. Brown A
    2. Rathore S
    3. Kimanius D
    4. Aibara S
    5. Bai XC
    6. Rorbach J
    7. Amunts A
    8. Ramakrishnan V

    (2017) Structures of the human mitochondrial ribosome in native states of assembly

    Nature Structural & Molecular Biology 24:866–869.

    https://doi.org/10.1038/nsmb.3464

    • PubMed
    • Google Scholar
15. 1. Brunger AT

    (2007) Version 1.2 of the crystallography and NMR system

    Nature Protocols 2:2728–2733.

    https://doi.org/10.1038/nprot.2007.406

    • PubMed
    • Google Scholar
16. 1. Cammack KA
    2. Wade HE

    (1965) The sedimentation behaviour of ribonuclease-active and -inactive ribosomes from Bacteria

    Biochemical Journal 96:671–680.

    https://doi.org/10.1042/bj0960671

    • PubMed
    • Google Scholar
17. 1. Capecchi MR

    (1967a) Polypeptide chain termination in vitro: isolation of a release factor

    PNAS 58:1144–1151.

    https://doi.org/10.1073/pnas.58.3.1144

    • PubMed
    • Google Scholar
18. 1. Capecchi MR

    (1967b) A rapid assay for polypeptide chain termination

    Biochemical and Biophysical Research Communications 28:773–778.

    https://doi.org/10.1016/0006-291X(67)90384-1

    • PubMed
    • Google Scholar
19. 1. Casy W
    2. Prater AR
    3. Cornish PV

    (2018) Operative binding of class I release factors and YaeJ stabilizes the ribosome in the nonrotated state

    Biochemistry 57:1954–1966.

    https://doi.org/10.1021/acs.biochem.7b00824

    • PubMed
    • Google Scholar
20. 1. Chapman MS

    (1995) Restrained real-space macromolecular atomic refinement using a new resolution-dependent electron-density function

    Acta Crystallographica Section a Foundations of Crystallography 51:69–80.

    https://doi.org/10.1107/S0108767394007130

    • Google Scholar
21. 1. Chen VB
    2. Arendall WB
    3. Headd JJ
    4. Keedy DA
    5. Immormino RM
    6. Kapral GJ
    7. Murray LW
    8. Richardson JS
    9. Richardson DC

    (2010) MolProbity: all-atom structure validation for macromolecular crystallography

    Acta Crystallographica Section D Biological Crystallography 66:12–21.

    https://doi.org/10.1107/S0907444909042073

    • PubMed
    • Google Scholar
22. 1. Chen Y
    2. Feng S
    3. Kumar V
    4. Ero R
    5. Gao YG

    (2013) Structure of EF-G-ribosome complex in a pretranslocation state

    Nature Structural & Molecular Biology 20:1077–1084.

    https://doi.org/10.1038/nsmb.2645

    • PubMed
    • Google Scholar
23. 1. Cornish PV
    2. Ermolenko DN
    3. Noller HF
    4. Ha T

    (2008) Spontaneous intersubunit rotation in single ribosomes

    Molecular Cell 30:578–588.

    https://doi.org/10.1016/j.molcel.2008.05.004

    • PubMed
    • Google Scholar
24. Software

    1. DeLano WL

    (2002)

    The PyMOL Molecular Graphics System

    DeLano Scientific, Palo Alto, CA, USA.
25. 1. Demo G
    2. Svidritskiy E
    3. Madireddy R
    4. Diaz-Avalos R
    5. Grant T
    6. Grigorieff N
    7. Sousa D
    8. Korostelev AA

    (2017) Mechanism of ribosome rescue by ArfA and RF2

    eLife 6:e23687.

    https://doi.org/10.7554/eLife.23687

    • PubMed
    • Google Scholar
26. 1. Dinçbas-Renqvist V
    2. Engström A
    3. Mora L
    4. Heurgué-Hamard V
    5. Buckingham R
    6. Ehrenberg M

    (2000) A post-translational modification in the GGQ motif of RF2 from Escherichia coli stimulates termination of translation

    The EMBO Journal 19:6900–6907.

    https://doi.org/10.1093/emboj/19.24.6900

    • PubMed
    • Google Scholar
27. 1. Dunkle JA
    2. Wang L
    3. Feldman MB
    4. Pulk A
    5. Chen VB
    6. Kapral GJ
    7. Noeske J
    8. Richardson JS
    9. Blanchard SC
    10. Cate JH

    (2011) Structures of the bacterial ribosome in classical and hybrid states of tRNA binding

    Science 332:981–984.

    https://doi.org/10.1126/science.1202692

    • PubMed
    • Google Scholar
28. 1. Dunkle JA
    2. Cate JH

    (2010) Ribosome structure and dynamics during translocation and termination

    Annual Review of Biophysics 39:227–244.

    https://doi.org/10.1146/annurev.biophys.37.032807.125954

    • PubMed
    • Google Scholar
29. 1. Ermolenko DN
    2. Majumdar ZK
    3. Hickerson RP
    4. Spiegel PC
    5. Clegg RM
    6. Noller HF

    (2007) Observation of intersubunit movement of the ribosome in solution using FRET

    Journal of Molecular Biology 370:530–540.

    https://doi.org/10.1016/j.jmb.2007.04.042

    • PubMed
    • Google Scholar
30. 1. Fischer N
    2. Konevega AL
    3. Wintermeyer W
    4. Rodnina MV
    5. Stark H

    (2010) Ribosome dynamics and tRNA movement by time-resolved electron cryomicroscopy

    Nature 466:329–333.

    https://doi.org/10.1038/nature09206

    • PubMed
    • Google Scholar
31. 1. Florin T
    2. Maracci C
    3. Graf M
    4. Karki P
    5. Klepacki D
    6. Berninghausen O
    7. Beckmann R
    8. Vázquez-Laslop N
    9. Wilson DN
    10. Rodnina MV
    11. Mankin AS

    (2017) An antimicrobial peptide that inhibits translation by trapping release factors on the ribosome

    Nature Structural & Molecular Biology 24:752–757.

    https://doi.org/10.1038/nsmb.3439

    • PubMed
    • Google Scholar
32. 1. Frank J
    2. Agrawal RK

    (2000) A ratchet-like inter-subunit reorganization of the ribosome during translocation

    Nature 406:318–322.

    https://doi.org/10.1038/35018597

    • PubMed
    • Google Scholar
33. 1. Freistroffer DV
    2. Pavlov MY
    3. MacDougall J
    4. Buckingham RH
    5. Ehrenberg M

    (1997) Release factor RF3 in e.coli accelerates the dissociation of release factors RF1 and RF2 from the ribosome in a GTP-dependent manner

    The EMBO Journal 16:4126–4133.

    https://doi.org/10.1093/emboj/16.13.4126

    • PubMed
    • Google Scholar
34. 1. Fu Z
    2. Kaledhonkar S
    3. Borg A
    4. Sun M
    5. Chen B
    6. Grassucci RA
    7. Ehrenberg M
    8. Frank J

    (2016) Key intermediates in ribosome recycling visualized by Time-Resolved cryoelectron microscopy

    Structure 24:2092–2101.

    https://doi.org/10.1016/j.str.2016.09.014

    • PubMed
    • Google Scholar
35. Preprint

    1. Fu Z
    2. Indrisiunaite G
    3. Kaledhonkar S
    4. Shah B
    5. Sun M
    6. Chen B
    7. Grassucci RA
    8. Ehrenberg M
    9. Frank J

    (2018) The structural basis for release factor activation during translation termination revealed by time-resolved cryogenic electron microscopy

    bioRxiv.

    https://doi.org/10.1016/j.bpj.2018.11.3090

    • Google Scholar
36. 1. Gao H
    2. Zhou Z
    3. Rawat U
    4. Huang C
    5. Bouakaz L
    6. Wang C
    7. Cheng Z
    8. Liu Y
    9. Zavialov A
    10. Gursky R
    11. Sanyal S
    12. Ehrenberg M
    13. Frank J
    14. Song H

    (2007) RF3 induces ribosomal conformational changes responsible for dissociation of class I release factors

    Cell 129:929–941.

    https://doi.org/10.1016/j.cell.2007.03.050

    • PubMed
    • Google Scholar
37. 1. Gao YG
    2. Selmer M
    3. Dunham CM
    4. Weixlbaumer A
    5. Kelley AC
    6. Ramakrishnan V

    (2009) The structure of the ribosome with elongation factor G trapped in the posttranslocational state

    Science 326:694–699.

    https://doi.org/10.1126/science.1179709

    • PubMed
    • Google Scholar
38. 1. Graf M
    2. Huter P
    3. Maracci C
    4. Peterek M
    5. Rodnina MV
    6. Wilson DN

    (2018) Visualization of translation termination intermediates trapped by the apidaecin 137 peptide during RF3-mediated recycling of RF1

    Nature Communications 9:3053.

    https://doi.org/10.1038/s41467-018-05465-1

    • PubMed
    • Google Scholar
39. 1. Grant T
    2. Rohou A
    3. Grigorieff N

    (2018) cisTEM, user-friendly software for single-particle image processing

    eLife 7:e35383.

    https://doi.org/10.7554/eLife.35383

    • PubMed
    • Google Scholar
40. 1. Grentzmann G
    2. Brechemier-Baey D
    3. Heurgue V
    4. Mora L
    5. Buckingham RH

    (1994) Localization and characterization of the gene encoding release factor RF3 in Escherichia coli

    PNAS 91:5848–5852.

    https://doi.org/10.1073/pnas.91.13.5848

    • PubMed
    • Google Scholar
41. 1. He SL
    2. Green R

    (2010) Visualization of codon-dependent conformational rearrangements during translation termination

    Nature Structural & Molecular Biology 17:465–470.

    https://doi.org/10.1038/nsmb.1766

    • PubMed
    • Google Scholar
42. 1. Hetrick B
    2. Lee K
    3. Joseph S

    (2009) Kinetics of stop Codon recognition by release factor 1

    Biochemistry 48:11178–11184.

    https://doi.org/10.1021/bi901577d

    • PubMed
    • Google Scholar
43. 1. Heymann JB
    2. Belnap DM

    (2007) Bsoft: image processing and molecular modeling for electron microscopy

    Journal of Structural Biology 157:3–18.

    https://doi.org/10.1016/j.jsb.2006.06.006

    • PubMed
    • Google Scholar
44. 1. Indrisiunaite G
    2. Pavlov MY
    3. Heurgué-Hamard V
    4. Ehrenberg M

    (2015) On the pH dependence of class-1 RF-dependent termination of mRNA translation

    Journal of Molecular Biology 427:1848–1860.

    https://doi.org/10.1016/j.jmb.2015.01.007

    • PubMed
    • Google Scholar
45. 1. James NR
    2. Brown A
    3. Gordiyenko Y
    4. Ramakrishnan V

    (2016) Translational termination without a stop Codon

    Science 354:1437–1440.

    https://doi.org/10.1126/science.aai9127

    • PubMed
    • Google Scholar
46. 1. JCSG

    (2005) Crystal structure of peptide chain release factor 1 (RF-1) (SMU.1085) from Streptococcus mutans at 2.34 A resolution

    JC.f.S. Genomics.

    https://doi.org/10.2210/pdb1ZBT/pdb

    • Google Scholar
47. 1. Jin H
    2. Kelley AC
    3. Loakes D
    4. Ramakrishnan V

    (2010) Structure of the 70S ribosome bound to release factor 2 and a substrate analog provides insights into catalysis of peptide release

    PNAS 107:8593–8598.

    https://doi.org/10.1073/pnas.1003995107

    • PubMed
    • Google Scholar
48. 1. Jin H
    2. Kelley AC
    3. Ramakrishnan V

    (2011) Crystal structure of the hybrid state of ribosome in complex with the Guanosine triphosphatase release factor 3

    PNAS 108:15798–15803.

    https://doi.org/10.1073/pnas.1112185108

    • PubMed
    • Google Scholar
49. 1. Julián P
    2. Konevega AL
    3. Scheres SH
    4. Lázaro M
    5. Gil D
    6. Wintermeyer W
    7. Rodnina MV
    8. Valle M

    (2008) Structure of ratcheted ribosomes with tRNAs in hybrid states

    PNAS 105:16924–16927.

    https://doi.org/10.1073/pnas.0809587105

    • PubMed
    • Google Scholar
50. 1. Klaholz BP
    2. Pape T
    3. Zavialov AV
    4. Myasnikov AG
    5. Orlova EV
    6. Vestergaard B
    7. Ehrenberg M
    8. van Heel M

    (2003) Structure of the Escherichia coli ribosomal termination complex with release factor 2

    Nature 421:90–94.

    https://doi.org/10.1038/nature01225

    • PubMed
    • Google Scholar
51. 1. Korostelev A
    2. Bertram R
    3. Chapman MS

    (2002) Simulated-annealing real-space refinement as a tool in model building

    Acta Crystallographica Section D Biological Crystallography 58:761–767.

    https://doi.org/10.1107/S0907444902003402

    • PubMed
    • Google Scholar
52. 1. Korostelev A
    2. Trakhanov S
    3. Laurberg M
    4. Noller HF

    (2006) Crystal structure of a 70S ribosome-tRNA complex reveals functional interactions and rearrangements

    Cell 126:1065–1077.

    https://doi.org/10.1016/j.cell.2006.08.032

    • PubMed
    • Google Scholar
53. 1. Korostelev A
    2. Asahara H
    3. Lancaster L
    4. Laurberg M
    5. Hirschi A
    6. Zhu J
    7. Trakhanov S
    8. Scott WG
    9. Noller HF

    (2008) Crystal structure of a translation termination complex formed with release factor RF2

    PNAS 105:19684–19689.

    https://doi.org/10.1073/pnas.0810953105

    • PubMed
    • Google Scholar
54. 1. Korostelev A
    2. Zhu J
    3. Asahara H
    4. Noller HF

    (2010) Recognition of the amber UAG stop Codon by release factor RF1

    The EMBO Journal 29:2577–2585.

    https://doi.org/10.1038/emboj.2010.139

    • PubMed
    • Google Scholar
55. 1. Korostelev AA

    (2011) Structural aspects of translation termination on the ribosome

    RNA 17:1409–1421.

    https://doi.org/10.1261/rna.2733411

    • PubMed
    • Google Scholar
56. 1. Koutmou KS
    2. McDonald ME
    3. Brunelle JL
    4. Green R

    (2014) RF3:gtp promotes rapid dissociation of the class 1 termination factor

    RNA 20:609–620.

    https://doi.org/10.1261/rna.042523.113

    • PubMed
    • Google Scholar
57. 1. Kremer JR
    2. Mastronarde DN
    3. McIntosh JR

    (1996) Computer visualization of three-dimensional image data using IMOD

    Journal of Structural Biology 116:71–76.

    https://doi.org/10.1006/jsbi.1996.0013

    • PubMed
    • Google Scholar
58. 1. Kuhlenkoetter S
    2. Wintermeyer W
    3. Rodnina MV

    (2011) Different substrate-dependent transition states in the active site of the ribosome

    Nature 476:351–354.

    https://doi.org/10.1038/nature10247

    • PubMed
    • Google Scholar
59. 1. Lancaster L
    2. Noller HF

    (2005) Involvement of 16S rRNA nucleotides G1338 and A1339 in discrimination of initiator tRNA

    Molecular Cell 20:623–632.

    https://doi.org/10.1016/j.molcel.2005.10.006

    • PubMed
    • Google Scholar
60. 1. Laurberg M
    2. Asahara H
    3. Korostelev A
    4. Zhu J
    5. Trakhanov S
    6. Noller HF

    (2008) Structural basis for translation termination on the 70S ribosome

    Nature 454:852–857.

    https://doi.org/10.1038/nature07115

    • PubMed
    • Google Scholar
61. 1. Leipe DD
    2. Wolf YI
    3. Koonin EV
    4. Aravind L

    (2002) Classification and evolution of P-loop GTPases and related ATPases

    Journal of Molecular Biology 317:41–72.

    https://doi.org/10.1006/jmbi.2001.5378

    • PubMed
    • Google Scholar
62. 1. Ling C
    2. Ermolenko DN

    (2016) Structural insights into ribosome translocation

    Wiley Interdisciplinary Reviews: RNA 7:620–636.

    https://doi.org/10.1002/wrna.1354

    • PubMed
    • Google Scholar
63. 1. Loveland AB
    2. Demo G
    3. Grigorieff N
    4. Korostelev AA

    (2017) Ensemble cryo-EM elucidates the mechanism of translation fidelity

    Nature 546:113–117.

    https://doi.org/10.1038/nature22397

    • PubMed
    • Google Scholar
64. 1. Loveland AB
    2. Korostelev AA

    (2018) Structural dynamics of protein S1 on the 70S ribosome visualized by ensemble cryo-EM

    Methods 137:55–66.

    https://doi.org/10.1016/j.ymeth.2017.12.004

    • PubMed
    • Google Scholar
65. 1. Lyumkis D
    2. Brilot AF
    3. Theobald DL
    4. Grigorieff N

    (2013) Likelihood-based classification of cryo-EM images using FREALIGN

    Journal of Structural Biology 183:377–388.

    https://doi.org/10.1016/j.jsb.2013.07.005

    • PubMed
    • Google Scholar
66. 1. Mastronarde DN

    (2005) Automated electron microscope tomography using robust prediction of specimen movements

    Journal of Structural Biology 152:36–51.

    https://doi.org/10.1016/j.jsb.2005.07.007

    • PubMed
    • Google Scholar
67. 1. Mikuni O
    2. Ito K
    3. Moffat J
    4. Matsumura K
    5. McCaughan K
    6. Nobukuni T
    7. Tate W
    8. Nakamura Y

    (1994) Identification of the prfC gene, which encodes peptide-chain-release factor 3 of Escherichia coli

    PNAS 91:5798–5802.

    https://doi.org/10.1073/pnas.91.13.5798

    • PubMed
    • Google Scholar
68. 1. Moazed D
    2. Noller HF

    (1989) Intermediate states in the movement of transfer RNA in the ribosome

    Nature 342:142–148.

    https://doi.org/10.1038/342142a0

    • PubMed
    • Google Scholar
69. 1. Nakamura Y
    2. Ito K
    3. Matsumura K
    4. Kawazu Y
    5. Ebihara K

    (1995) Regulation of translation termination: conserved structural motifs in bacterial and eukaryotic polypeptide release factors

    Biochemistry and Cell Biology 73:1113–1122.

    https://doi.org/10.1139/o95-120

    • PubMed
    • Google Scholar
70. 1. Nichols RJ
    2. Sen S
    3. Choo YJ
    4. Beltrao P
    5. Zietek M
    6. Chaba R
    7. Lee S
    8. Kazmierczak KM
    9. Lee KJ
    10. Wong A
    11. Shales M
    12. Lovett S
    13. Winkler ME
    14. Krogan NJ
    15. Typas A
    16. Gross CA

    (2011) Phenotypic landscape of a bacterial cell

    Cell 144:143–156.

    https://doi.org/10.1016/j.cell.2010.11.052

    • Google Scholar
71. 1. Noller HF
    2. Lancaster L
    3. Zhou J
    4. Mohan S

    (2017) The ribosome moves: rna mechanics and translocation

    Nature Structural & Molecular Biology 24:1021–1027.

    https://doi.org/10.1038/nsmb.3505

    • PubMed
    • Google Scholar
72. 1. O'Connor M

    (2015) Interactions of release factor RF3 with the translation machinery

    Molecular Genetics and Genomics 290:1335–1344.

    https://doi.org/10.1007/s00438-015-0994-x

    • PubMed
    • Google Scholar
73. 1. Pai RD
    2. Zhang W
    3. Schuwirth BS
    4. Hirokawa G
    5. Kaji H
    6. Kaji A
    7. Cate JH

    (2008) Structural insights into ribosome recycling factor interactions with the 70S ribosome

    Journal of Molecular Biology 376:1334–1347.

    https://doi.org/10.1016/j.jmb.2007.12.048

    • PubMed
    • Google Scholar
74. 1. Pallesen J
    2. Hashem Y
    3. Korkmaz G
    4. Koripella RK
    5. Huang C
    6. Ehrenberg M
    7. Sanyal S
    8. Frank J

    (2013) Cryo-EM visualization of the ribosome in termination complex with apo-RF3 and RF1

    eLife 2:e00411.

    https://doi.org/10.7554/eLife.00411

    • PubMed
    • Google Scholar
75. 1. Petry S
    2. Brodersen DE
    3. Murphy FV
    4. Dunham CM
    5. Selmer M
    6. Tarry MJ
    7. Kelley AC
    8. Ramakrishnan V

    (2005) Crystal structures of the ribosome in complex with release factors RF1 and RF2 bound to a cognate stop Codon

    Cell 123:1255–1266.

    https://doi.org/10.1016/j.cell.2005.09.039

    • Google Scholar
76. 1. Pettersen EF
    2. Goddard TD
    3. Huang CC
    4. Couch GS
    5. Greenblatt DM
    6. Meng EC
    7. Ferrin TE

    (2004) UCSF chimera--a visualization system for exploratory research and analysis

    Journal of Computational Chemistry 25:1605–1612.

    https://doi.org/10.1002/jcc.20084

    • PubMed
    • Google Scholar
77. 1. Pierson WE
    2. Hoffer ED
    3. Keedy HE
    4. Simms CL
    5. Dunham CM
    6. Zaher HS

    (2016) Uniformity of peptide release is maintained by methylation of release factors

    Cell Reports 17:11–18.

    https://doi.org/10.1016/j.celrep.2016.08.085

    • PubMed
    • Google Scholar
78. 1. Polacek N
    2. Gomez MJ
    3. Ito K
    4. Xiong L
    5. Nakamura Y
    6. Mankin A

    (2003) The critical role of the universally conserved A2602 of 23S ribosomal RNA in the release of the nascent peptide during translation termination

    Molecular Cell 11:103–112.

    https://doi.org/10.1016/S1097-2765(02)00825-0

    • PubMed
    • Google Scholar
79. 1. Polikanov YS
    2. Steitz TA
    3. Innis CA

    (2014) A proton wire to couple aminoacyl-tRNA accommodation and peptide-bond formation on the ribosome

    Nature Structural & Molecular Biology 21:787–793.

    https://doi.org/10.1038/nsmb.2871

    • Google Scholar
80. 1. Prabhakar A
    2. Capece MC
    3. Petrov A
    4. Choi J
    5. Puglisi JD

    (2017) Post-termination ribosome intermediate acts as the gateway to ribosome recycling

    Cell Reports 20:161–172.

    https://doi.org/10.1016/j.celrep.2017.06.028

    • PubMed
    • Google Scholar
81. 1. Pulk A
    2. Cate JH

    (2013) Control of ribosomal subunit rotation by elongation factor G

    Science 340:1235970.

    https://doi.org/10.1126/science.1235970

    • PubMed
    • Google Scholar
82. 1. Ramakrishnan V

    (2011)

    Ribosomes: Structure, Function, and Dynamics

    Structural Studies on Decoding, Termination and Translocation in the Bacterial Ribosome, Ribosomes: Structure, Function, and Dynamics, Springer.

    • Google Scholar
83. 1. Ratje AH
    2. Loerke J
    3. Mikolajka A
    4. Brünner M
    5. Hildebrand PW
    6. Starosta AL
    7. Dönhöfer A
    8. Connell SR
    9. Fucini P
    10. Mielke T
    11. Whitford PC
    12. Onuchic JN
    13. Yu Y
    14. Sanbonmatsu KY
    15. Hartmann RK
    16. Penczek PA
    17. Wilson DN
    18. Spahn CM

    (2010) Head swivel on the ribosome facilitates translocation by means of intra-subunit tRNA hybrid sites

    Nature 468:713–716.

    https://doi.org/10.1038/nature09547

    • PubMed
    • Google Scholar
84. 1. Rawat UB
    2. Zavialov AV
    3. Sengupta J
    4. Valle M
    5. Grassucci RA
    6. Linde J
    7. Vestergaard B
    8. Ehrenberg M
    9. Frank J

    (2003) A cryo-electron microscopic study of ribosome-bound termination factor RF2

    Nature 421:87–90.

    https://doi.org/10.1038/nature01224

    • PubMed
    • Google Scholar
85. 1. Rawat U
    2. Gao H
    3. Zavialov A
    4. Gursky R
    5. Ehrenberg M
    6. Frank J

    (2006) Interactions of the release factor RF1 with the ribosome as revealed by cryo-EM

    Journal of Molecular Biology 357:1144–1153.

    https://doi.org/10.1016/j.jmb.2006.01.038

    • PubMed
    • Google Scholar
86. 1. Rodnina MV

    (2018) Translation in prokaryotes

    Cold Spring Harbor Perspectives in Biology 10:a032664.

    https://doi.org/10.1101/cshperspect.a032664

    • Google Scholar
87. 1. Santos N
    2. Zhu J
    3. Donohue JP
    4. Korostelev AA
    5. Noller HF

    (2013) Crystal structure of the 70S ribosome bound with the Q253P mutant form of release factor RF2

    Structure 21:1258–1263.

    https://doi.org/10.1016/j.str.2013.04.028

    • PubMed
    • Google Scholar
88. 1. Scheres SH
    2. Gao H
    3. Valle M
    4. Herman GT
    5. Eggermont PP
    6. Frank J
    7. Carazo JM

    (2007) Disentangling conformational states of macromolecules in 3D-EM through likelihood optimization

    Nature Methods 4:27–29.

    https://doi.org/10.1038/nmeth992

    • PubMed
    • Google Scholar
89. 1. Scheres SH

    (2010) Classification of structural heterogeneity by maximum-likelihood methods

    Methods in Enzymology 482:295–320.

    https://doi.org/10.1016/S0076-6879(10)82012-9

    • PubMed
    • Google Scholar
90. 1. Scolnick E
    2. Tompkins R
    3. Caskey T
    4. Nirenberg M

    (1968) Release factors differing in specificity for terminator codons

    PNAS 61:768–774.

    https://doi.org/10.1073/pnas.61.2.768

    • PubMed
    • Google Scholar
91. 1. Selmer M
    2. Dunham CM
    3. Murphy FV
    4. Weixlbaumer A
    5. Petry S
    6. Kelley AC
    7. Weir JR
    8. Ramakrishnan V

    (2006) Structure of the 70S ribosome complexed with mRNA and tRNA

    Science 313:1935–1942.

    https://doi.org/10.1126/science.1131127

    • PubMed
    • Google Scholar
92. 1. Shi X
    2. Joseph S

    (2016) Mechanism of translation termination: rf1 dissociation follows dissociation of RF3 from the ribosome

    Biochemistry 55:6344–6354.

    https://doi.org/10.1021/acs.biochem.6b00921

    • PubMed
    • Google Scholar
93. 1. Shin DH
    2. Brandsen J
    3. Jancarik J
    4. Yokota H
    5. Kim R
    6. Kim SH

    (2004) Structural analyses of peptide release factor 1 from Thermotoga maritima reveal domain flexibility required for its interaction with the ribosome

    Journal of Molecular Biology 341:227–239.

    https://doi.org/10.1016/j.jmb.2004.05.055

    • PubMed
    • Google Scholar
94. 1. Sternberg SH
    2. Fei J
    3. Prywes N
    4. McGrath KA
    5. Gonzalez RL

    (2009) Translation factors direct intrinsic ribosome dynamics during translation termination and ribosome recycling

    Nature Structural & Molecular Biology 16:861–868.

    https://doi.org/10.1038/nsmb.1622

    • PubMed
    • Google Scholar
95. 1. Svidritskiy E
    2. Brilot AF
    3. Koh CS
    4. Grigorieff N
    5. Korostelev AA

    (2014) Structures of yeast 80S ribosome-tRNA complexes in the rotated and nonrotated conformations

    Structure 22:1210–1218.

    https://doi.org/10.1016/j.str.2014.06.003

    • PubMed
    • Google Scholar
96. 1. Svidritskiy E
    2. Madireddy R
    3. Korostelev AA

    (2016) Structural basis for translation termination on a pseudouridylated stop Codon

    Journal of Molecular Biology 428:2228–2236.

    https://doi.org/10.1016/j.jmb.2016.04.018

    • PubMed
    • Google Scholar
97. 1. Svidritskiy E
    2. Demo G
    3. Korostelev AA

    (2018) Mechanism of premature translation termination on a sense Codon

    Journal of Biological Chemistry 293:12472–12479.

    https://doi.org/10.1074/jbc.AW118.003232

    • PubMed
    • Google Scholar
98. 1. Svidritskiy E
    2. Korostelev AA

    (2018a) Conformational control of translation termination on the 70S ribosome

    Structure 26:821–828.

    https://doi.org/10.1016/j.str.2018.04.001

    • PubMed
    • Google Scholar
99. 1. Svidritskiy E
    2. Korostelev AA

    (2018b) Mechanism of inhibition of translation termination by blasticidin S

    Journal of Molecular Biology 430:591–593.

    https://doi.org/10.1016/j.jmb.2018.01.007

    • PubMed
    • Google Scholar
100. 1. Tompkins RK
     2. Scolnick EM
     3. Caskey CT

     (1970) Peptide chain termination. VII. The ribosomal and release factor requirements for peptide release

     PNAS 65:702–708.

     https://doi.org/10.1073/pnas.65.3.702

     • PubMed
     • Google Scholar
101. 1. Tourigny DS
     2. Fernández IS
     3. Kelley AC
     4. Ramakrishnan V

     (2013) Elongation factor G bound to the ribosome in an intermediate state of translocation

     Science 340:1235490.

     https://doi.org/10.1126/science.1235490

     • PubMed
     • Google Scholar
102. 1. Trappl K
     2. Joseph S

     (2016) Ribosome induces a closed to open conformational change in release factor 1

     Journal of Molecular Biology 428:1333–1344.

     https://doi.org/10.1016/j.jmb.2016.01.021

     • PubMed
     • Google Scholar
103. 1. Vestergaard B
     2. Van LB
     3. Andersen GR
     4. Nyborg J
     5. Buckingham RH
     6. Kjeldgaard M

     (2001) Bacterial polypeptide release factor RF2 is structurally distinct from eukaryotic eRF1

     Molecular Cell 8:1375–1382.

     https://doi.org/10.1016/S1097-2765(01)00415-4

     • PubMed
     • Google Scholar
104. 1. Weixlbaumer A
     2. Jin H
     3. Neubauer C
     4. Voorhees RM
     5. Petry S
     6. Kelley AC
     7. Ramakrishnan V

     (2008) Insights into translational termination from the structure of RF2 bound to the ribosome

     Science 322:953–956.

     https://doi.org/10.1126/science.1164840

     • PubMed
     • Google Scholar
105. 1. Youngman EM
     2. McDonald ME
     3. Green R

     (2008) Peptide release on the ribosome: mechanism and implications for translational control

     Annual Review of Microbiology 62:353–373.

     https://doi.org/10.1146/annurev.micro.61.080706.093323

     • PubMed
     • Google Scholar
106. 1. Zavialov AV
     2. Mora L
     3. Buckingham RH
     4. Ehrenberg M

     (2002) Release of peptide promoted by the GGQ motif of class 1 release factors regulates the GTPase activity of RF3

     Molecular Cell 10:789–798.

     https://doi.org/10.1016/S1097-2765(02)00691-3

     • PubMed
     • Google Scholar
107. 1. Zeng F
     2. Jin H

     (2018) Conformation of methylated GGQ in the peptidyl transferase center during translation termination

     Scientific Reports 8:2349.

     https://doi.org/10.1038/s41598-018-20107-8

     • PubMed
     • Google Scholar
108. 1. Zhou J
     2. Lancaster L
     3. Trakhanov S
     4. Noller HF

     (2012) Crystal structure of release factor RF3 trapped in the GTP state on a rotated conformation of the ribosome

     RNA 18:230–240.

     https://doi.org/10.1261/rna.031187.111

     • PubMed
     • Google Scholar
109. 1. Zhou J
     2. Lancaster L
     3. Donohue JP
     4. Noller HF

     (2013) Crystal structures of EF-G-ribosome complexes trapped in intermediate states of translocation

     Science 340:1236086.

     https://doi.org/10.1126/science.1236086

     • PubMed
     • Google Scholar
110. 1. Zoldák G
     2. Redecke L
     3. Svergun DI
     4. Konarev PV
     5. Voertler CS
     6. Dobbek H
     7. Sedlák E
     8. Sprinzl M

     (2007) Release factors 2 from Escherichia coli and Thermus thermophilus: structural, spectroscopic and microcalorimetric studies

     Nucleic Acids Research 35:1343–1353.

     https://doi.org/10.1093/nar/gkl696

     • PubMed
     • Google Scholar

Author details

1. Egor Svidritskiy

   RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, United States

   Contribution

   Conceptualization, Data curation, Formal analysis, Validation, Investigation, Visualization, Methodology, Writing—review and editing, Assembled the ribosome complexes, Performed biochemistry, Collected and processed cryo-EM data, Modeled and refined structural models

   Competing interests

   No competing interests declared

2. Gabriel Demo

   RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, United States

   Contribution

   Data curation, Formal analysis, Investigation, Methodology, Purified ribosome subunits and RF2, Assisted with structure refinements

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0002-5472-9249

3. Anna B Loveland

   RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, United States

   Contribution

   Formal analysis, Methodology, Assisted with data processing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0001-9172-7747

4. Chen Xu

   Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, United States

   Contribution

   Methodology, Implemented multiple-shot (per hole) data acquisition, Optimized data collection

   Competing interests

   No competing interests declared

5. Andrei A Korostelev

   1. RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, United States
   2. Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, United States

   Contribution

   Conceptualization, Resources, Formal analysis, Supervision, Funding acquisition, Validation, Investigation, Writing—original draft, Project administration, Writing—review and editing, Supervised the project, Assisted with modeling and structure refinements, Wrote the manuscript with input from co-authors

   For correspondence

   andrei.korostelev@umassmed.edu

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0003-1588-717X

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