Structure and Binding Mode of a Ribosome Recycling Factor (RRF) from Mesophilic Bacterium

Nakano, Hiroaki; Yoshida, Takuya; Uchiyama, Susumu; Kawachi, Masako; Matsuo, Hitomi; Kato, Takayuki; Ohshima, Atsushi; Yamaichi, Yoshiharu; Honda, Takeshi; Kato, Hiroaki; Yamagata, Yuriko; Ohkubo, Tadayasu; Kobayashi, Yuji

doi:10.1074/jbc.m208098200

Cited by 38 publications

(57 citation statements)

References 41 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The L-shape of the RRF density in the cryo-EM map ( Fig. 1) is consistent with crystal (9,11,13) and solution (12) structures of RRF from various species. Independent docking of all these atomic structures into the EM density shows that the density is best explained by the crystallographic structure of the Thermotoga maritima RRF (Fig.…”

Section: Comparison Of Atomic Structures Of Rrf With Corresponding Crsupporting

confidence: 59%

“…Atomic structures of RRF determined from five different species, including Escherichia coli, show that it is comprised of two structural domains: domain I, consisting of three long ␣-helix bundles, and the smaller domain II, which is an ␣͞␤ motif (9)(10)(11)(12)(13). Different orientations of domain II in these structures have been attributed to interdomain flexibility, which is thought to be necessary for RRF to function on the ribosome (12).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Visualization of ribosome-recycling factor on the Escherichia coli 70S ribosome: Functional implications

Agrawal

Sharma

Kiel

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

148

170

View full text Add to dashboard Cite

After the termination step of protein synthesis, a deacylated tRNA and mRNA remain associated with the ribosome. The ribosomerecycling factor (RRF), together with elongation factor G (EF-G), disassembles this posttermination complex into mRNA, tRNA, and the ribosome. We have obtained a three-dimensional cryo-electron microscopic map of a complex of the Escherichia coli 70S ribosome and RRF. We find that RRF interacts mainly with the segments of the large ribosomal subunit's (50S) rRNA helices that are involved in the formation of two central intersubunit bridges, B2a and B3. The binding of RRF induces considerable conformational changes in some of the functional domains of the ribosome. As compared to its binding position derived previously by hydroxyl radical probing study, we find that RRF binds further inside the intersubunit space of the ribosome such that the tip of its domain I is shifted (by Ϸ13 Å) toward protein L5 within the central protuberance of the 50S subunit, and domain II is oriented more toward the small ribosomal subunit (30S). Overlapping binding sites of RRF, EF-G, and the P-site tRNA suggest that the binding of EF-G would trigger the removal of deacylated tRNA from the P site by moving RRF toward the ribosomal E site, and subsequent removal of mRNA may be induced by a shift in the position of 16S rRNA helix 44, which harbors part of the mRNA. R ibosomes are responsible for translating genetic information carried by mRNAs into specific sequences of amino acids. Translation on the ribosome comprises of four main steps: (i) initiation, (ii) elongation, (iii) termination, and (iv) recycling. Recent advancements in structural studies of the translational machinery have helped elucidate binding positions and functions of various translational factors involved in various stages of initiation (1, 2), elongation (3, 4), and termination (5-7). The fourth step of translation requires binding of a dedicated protein factor, the ribosome-recycling factor (RRF), which in conjunction with elongation factor G (EF-G) helps removing the mRNA and last deacylated tRNA from the ribosome (see ref. 8).Atomic structures of RRF determined from five different species, including Escherichia coli, show that it is comprised of two structural domains: domain I, consisting of three long ␣-helix bundles, and the smaller domain II, which is an ␣͞␤ motif (9-13). Different orientations of domain II in these structures have been attributed to interdomain flexibility, which is thought to be necessary for RRF to function on the ribosome (12).The overall match in dimensions between RRF and tRNA (9) prompted the proposal of structural and functional molecular mimicry between the two molecules (9). In a recent study (14) using the hydroxyl radical probing (HRP) method, the orientation of RRF on the ribosome was derived. This study did not support the idea of direct molecular mimicry of tRNA by RRF, because the derived binding position of RRF was quite different from that one would expect based on structural mimicry. The inferred bin...

show abstract

Section: Comparison Of Atomic Structures Of Rrf With Corresponding Crsupporting

confidence: 59%

mentioning

confidence: 99%

Visualization of ribosome-recycling factor on the Escherichia coli 70S ribosome: Functional implications

Agrawal

Sharma

Kiel

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

148

170

View full text Add to dashboard Cite

show abstract

“…Crystal and solution structures of RRF from several different organisms show that it is composed of two domains that adopt an "L" configuration [13][14][15][16][17][18][19] . Domain I of RRF is composed of a three-helix bundle ( Figure 2) that interacts with the large subunit of the ribosome 20-22 .…”

Section: Introductionmentioning

confidence: 99%

Structural Insights into Ribosome Recycling Factor Interactions with the 70S Ribosome

Pai

Zhang

Schuwirth

et al. 2008

Journal of Molecular Biology

View full text Add to dashboard Cite

SUMMARYAt the end of translation in bacteria, ribosome recycling factor (RRF) is used together with Elongation Factor G (EF-G) to recycle the 30S and 50S ribosomal subunits for the next round of translation. In x-ray crystal structures of RRF with the Escherichia coli 70S ribosome, RRF binds to the large ribosomal subunit in the cleft that contains the peptidyl transferase center (PTC). Upon binding of either E. coli or T. thermophilus RRF to the E. coli ribosome, the tip of ribosomal RNA helix H69 in the large subunit moves away from the small subunit toward RRF by 8 Å, thereby disrupting a key contact between the small and large ribosomal subunits, termed bridge B2a. In the ribosome crystals, the ability of RRF to destabilize bridge B2a is influenced by crystal packing forces. Movement of H69 involves an ordered to disordered transition upon binding of RRF to the ribosome. The disruption of bridge B2a upon RRF binding to the ribosome seen in the present structures reveals one of the key roles that RRF plays in ribosome recycling, the dissociation of 70S ribosomes into subunits. The structures also reveal contacts between Domain II of RRF and protein S12 in the 30S subunit that may also play a role in ribosome recycling.

show abstract

“…The three ␣-helices are arranged with a very rare right-handed twist. We found that, by using the DALI server (19,20), this helical arrangement has only been seen before for domain I of the ribosome recycling factor from different prokaryotic organisms (21)(22)(23)(24)(25).…”

Section: Conformational Heterogeneity Of Ahsp-thementioning

confidence: 98%

NMR Structure of the α-Hemoglobin Stabilizing Protein

Santiveri

Pérez‐Cañadillas

Vadivelu

et al. 2004

Journal of Biological Chemistry

View full text Add to dashboard Cite

The structure of ␣-hemoglobin stabilizing protein (AHSP), a molecular chaperone for free ␣-hemoglobin, has been determined using NMR spectroscopy. The protein native state shows conformational heterogeneity attributable to the isomerization of the peptide bond preceding a conserved proline residue. The two equally populated cis and trans forms both adopt an elongated antiparallel three ␣-helix bundle fold but display major differences in the loop between the first two helices and at the C terminus of helix 3. Proline to alanine single point mutation of the residue Pro-30 prevents the cis/ trans isomerization. The structure of the P30A mutant is similar to the structure of the trans form of AHSP in the loop 1 region. Both the wild-type AHSP and the P30A mutant bind to ␣-hemoglobin, and the wild-type conformational heterogeneity is quenched upon complex formation, suggesting that just one conformation is the active form. Changes in chemical shift observed upon complex formation identify a binding interface comprising the C terminus of helix 1, the loop 1, and the N terminus of helix 2, with the exposed residues Phe-47 and Tyr-51 being attractive targets for molecular recognition. The characteristics of this interface suggest that AHSP binds at the intradimer ␣ 1 ␤ 1 interface in tetrameric HbA.␣-Hemoglobin stabilizing protein (AHSP), 1 previously named EDRF (erythroid differentiation-related factor), was originally identified as a protein that showed a decrease in expression in the hematopoietic tissues of rodents and cattle infected with transmissible spongiform encephalopathies (1). EDRF was renamed AHSP after it was found that it binds specifically to ␣-hemoglobin in vitro (2). AHSP knock-out mice show erythrocyte abnormalities similar to those observed in ␤-thalassemia (2), and loss of AHSP aggravates the ␤-thalassemia phenotype in mice (38). ␤-Thalassemia is characterized by a reduction in the synthesis of ␤-globin chains. Consequently, the ␣-chains are in excess, and some of them precipitate, leading to defective red blood cells that determine the pathophysiology of ␤-thalassemia syndromes. The in vivo and in vitro data therefore both suggest that AHSP acts as a molecular chaperone for free ␣-hemoglobin (2, 3).Both AHSP and ␣-hemoglobin are monomeric in solution and bind each other with an association constant of 1 ϫ 10 7 M Ϫ1forming a complex with a 1:1 stoichiometry (4). The aim of the present study was to structurally characterize the AHSP-␣-hemoglobin interaction to understand the molecular basis of the AHSP function and its role as a hemoglobin chaperone. We have demonstrated by NMR that the AHSP native state is conformationally heterogeneous in solution and have identified the dynamic source of this behavior as a cis/trans isomerization of residue Pro-30. The slow time scale of this phenomenon allowed us to determine the high resolution three-dimensional structure of the two conformations of the wild-type AHSP under conditions where both conformations are equally populated. We were able to quench the c...

show abstract

Structure and Binding Mode of a Ribosome Recycling Factor (RRF) from Mesophilic Bacterium

Cited by 38 publications

References 41 publications

Visualization of ribosome-recycling factor on the Escherichia coli 70S ribosome: Functional implications

Visualization of ribosome-recycling factor on the Escherichia coli 70S ribosome: Functional implications

Structural Insights into Ribosome Recycling Factor Interactions with the 70S Ribosome

NMR Structure of the α-Hemoglobin Stabilizing Protein

Contact Info

Product

Resources

About