GTC
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE ADVANCED SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Genes to Cells (2004) 9, 271-278. doi:10.1111/j.1356-9597.2004.00723.x
© 2004 Blackwell Publishing or its licensors
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Yoshida, H.
Right arrow Articles by Wada, A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Yoshida, H.
Right arrow Articles by Wada, A.

RMF inactivates ribosomes by covering the peptidyl transferase centre and entrance of peptide exit tunnel

Hideji Yoshida1,*, Hiroshi Yamamoto2, Toshio Uchiumi2 and Akira Wada1

1 Department of Physics, Osaka Medical College, Takatsuki, Osaka 569-8686, Japan
2 Institute of High Polymer Research, Faculty of Textile Science and Technology, Shinshu University, Ueda 386-8567, Japan


   Abstract
Top
 Abstract
Introduction
 Results
 Discussion
 Experimental procedures
 References
 
In gram-negative bacteria such as Escherichia coli, protein synthesis is suppressed by the formation of 100S ribosomes under stress conditions. The 100S ribosome, a dimer of 70S ribosomes, is formed by ribosome modulation factor (RMF) binding to the 70S ribosomes. During the stationary phase, most of the 70S ribosomes turn to 100S ribosomes, which have lost translational activity. This 100S formation is called the hibernation process in the ribosome cycle of the stationary phase. If stationary phase cells are transferred to fresh medium, the 100S ribosomes immediately go back to active 70S ribosomes, showing that inactive 100S {leftrightarrow} active 70S interconversion is a major system regulating translation activity in stationary phase cells. To elucidate the mechanisms of translational inactivation, the binding sites of RMF on 23S rRNA in 100S ribosome of E. coli were examined by a chemical probing method using dimethyl sulphate (DMS). As the results, the nine bases in 23S rRNA were protected from DMS modifications and the modification of one base was enhanced. Interestingly A2451 is included among the protected bases, which is thought to be directly involved in peptidyl transferase activity. We conclude that RMF inactivates ribosomes by covering the peptidyl transferase (PTase) centre and the entrance of peptide exit tunnel. It is surprising that the cell itself produces a protein that seems to inhibit protein synthesis in a similar manner to antibiotics and that it can reversibly bind to and release from the ribosome in response to environmental conditions.


   Introduction
Top
 Abstract
 Introduction
Results
 Discussion
 Experimental procedures
 References
 
Cells under stress conditions, such as starvation, suppress protein synthesis using different strategies. The repression of ribosomal biosynthesis has been known to exist among many organisms (Mizuta & Warner 1994). In eukaryotes, it is known that phosphorylation of translational initiation factor-2 (eIF-2) is an adaptive mechanism for down-regulating protein synthesis under stress conditions (Hinnebusch 1994). In prokaryotes such as enterobacteria including Escherichia coli, the protein synthesis is suppressed by the formation of 100S ribosomes (Wada et al. 1990). The 100S ribosome, a dimer of 70S ribosomes, is formed by ribosome modulation factor (RMF) binding to the 70S ribosomes. The RMF is a basic (pI = 11.3) and small (Mr = 6507) protein, and its expression remarkably increases during the transition from the exponential to the stationary growth phase. It was reported that the rmf gene is induced by ppGpp and transcribed during the stationary phase (Izutsu et al. 2001a). Many proteins expressed in the stationary phase increase and decrease, existing for a limited period, but ribosomal binding proteins, RMF, YfiA, YhbH (Maki et al. 2000) and SRA (Izutsu et al. 2001b), exist throughout the stationary phase. It is suggested that these four proteins play important roles for survival during the stationary phase.

The 100S ribosomes have no translational activity and are formed during the stationary phase (Wada et al. 1995). When stationary phase cells are transferred to rich nutritious culture medium, RMF is immediately released from the 100S ribosomes, and the 100S ribosomes dissociate back into the 70S ribosomes. This process is very quick and completed within two minutes. After the process, the cells can reinitiate the protein synthesis and the proliferation within six minutes (Wada 1998). These phenomena indicate that an interconversion system between 70S and 100S ribosomes is a response to the transition of growth phases. During the exponential phase, in order to synthesize proteins, the ribosomes proceed step by step in the ribosome cycle that mainly consists of initiation, elongation, termination and recycling stages. During the stationary phase many ribosomes are changed to 100S ribosomes after the recycling stage. This 100S formation is called the hibernation stage (Yoshida et al. 2002). It is suggested that the 100S formation has roles of protection from ribosomal degradation in addition to suppression of protein synthesis, because the lifetime of RMF disruptants (E. coli) is shorter than that of wild strains (Yamagishi et al. 1993).

The above-mentioned facts raise the question of how RMF forms the 100S ribosomes and deprives the ribosomal particles of the translational activity. To elucidate these subjects, we have examined the binding site of RMF on the E. coli ribosome by protein-protein crosslinking using 2-iminothiolane in the previous studies (Yoshida et al. 2002). The results indicate that RMF is crosslinked with ribosomal proteins L2, L13 and S13, and we derived the possibility that RMF binds to a region at or near the peptidyl transferase (PTase) centre, because these ribosomal proteins exist around the PTase centre. This interpretation can explain the fact that the 100S ribosome has no translational activity. X-ray crystallographic analysis of ribosome indicates that no ribosomal protein is found in the PTase centre (Nissen et al. 2000). Therefore, it is predicted that RMF directly binds to the 23S rRNA. A knowledge of the interaction between RMF and the 23S rRNA is very valuable for understanding the role of RMF, because the rRNA carries out many ribosomal functions. In this study, we examined the RMF binding sites on the E. coli 23S rRNA by a chemical probing method using dimethyl sulphate (DMS). As a result, we found 10 ribonucleotides whose modifications are affected by RMF in the presence of DMS. Nine nucleotides among them are protected from DMS modification, and their positions are basically consistent with those of a crosslinking study. Interestingly the binding sites of RMF demonstrated in this study overlap with those of tRNAs and some antibiotics. Moreover, some of the detected nucleotides are crucial to the PTase activity and the evacuation of synthesized peptides from the ribosome.


   Results
Top
 Abstract
 Introduction
 Results
Discussion
 Experimental procedures
 References
 
In this study, the binding sites of RMF on E. coli 23S rRNA were examined by using DMS chemical probing. The previous studies of protein-protein crosslinking using 2-iminothiolane elucidated that RMF crosslinks to ribosomal proteins L2 and L13 on the 50S subunit, suggesting that RMF is located near the PTase centre (Yoshida et al. 2002). On the basis of this information, we have analysed the extensive regions of 23S rRNA including the PTase centre.

The formation of 100S ribosomes in vitro was performed by incubation of the RMF-free ribosomes (High-salt washed ribosomes: HSR, Fig. 1a) isolated from cells in stationary phase with His-tagged RMF in DMS reaction buffer (100 mM KCl, 15 mM MgCl2, 50 mM K-cacodylate at pH 7.2) as shown in Fig. 1c. Efficiency of the 100S ribosome formation in DMS reaction buffer detected by the sucrose-gradient sedimentation analysis was comparable to that formed in the association buffer (100 mM CH3COONH4, 15 mM (CH3COO)2Mg, and 20 mM Tris-HCl at pH 7.6). We also used the 100S ribosomes containing RMF formed in vivo as the RMF-containing 100S sample (Fig. 1b). The 100S ribosomes formed in vitro or in vivo were exposed to DMS, which modifies adenosine and cytidine of the rRNA at their N1 and N3 position, respectively. The modified rRNA bases were readily detected by primer extension analysis. We searched the regions of A920-G1107, A1918-A2090 and C2300-A2710 by using nine primers (Table 1) and performed the experiments at least more than three times for each of the 100S ribosomes formed in vitro and in vivo. The RMF-protection patterns were basically same between the two 100S ribosome samples, although there were some minor differences in the protection between the samples. It is therefore likely that there is no significant difference in structural nature between the 100S ribosomes formed in vitro and in vivo, despite the inefficiency in formation of 100S ribosomes with the His-tagged RMF in vitro compare to those formed with natural RMF as shown in Fig. 1b,c.



View larger version (14K):
[in this window]
[in a new window]
  		Figure 1  100S ribosomes in DMS reaction buffer. The high-salt washed ribosomes (a) and crude ribosomes (b) were prepared in DMS reaction buffer from the cells in the stationary phase. The 100S ribosomes formed in vitro (c) were prepared by adding the His-tagged RMF to HSR in DMS reaction buffer. The ribosomes were centrifuged on 5–20% linear sucrose density gradients at 40 000 r.p.m. (285 000 g) for 80 min.

 

View this table:
[in this window]
[in a new window]
  		Table 1  Primers used in extension reaction (E. coli numbering)
 
Figure 2 shows effects of RMF binding detected in the DMS-footprinting. Only major effects, which are reproducible and common to both the 100S samples, are shown. The four lanes on the left are dideoxy sequencing lanes. The protected and enhanced bases are judged in the fifth and sixth lanes from the left. DMS modification makes reverse transcriptase stop at the position immediately preceding the modified base. Considering these points, we observed 10 bases that were protected from DMS modifications and two bases whose modifications were enhanced by RMF binding. It is assumed that the protected bases escape the DMS modification by RMF binding, while the enhanced bases are exposed to DMS by conformational changes. This idea indicates a possibility that protected bases escaped the modification not only by physical contacts with RMF but also by conformational changes. Therefore, in order to discuss the RMF binding sites from the results of chemical probing, we examined a wide range of bases in the 23S rRNA and looked more closely at strongly modified bases. The protected bases were A1918, A1927, A1936, U1940, A2058, A2059, A2062, C2394, A2439 and A2451 (E. coli numbering). The enhanced bases were A2432 and U2571. The bases U1940 and U2571 were apparently protected and enhanced, respectively. The stop signals at U bases are unexpected, because uridine bases are not usual targets of DMS. These may be caused by a reason other than DMS modification. Therefore, we considered that nine bases, except U, were protected from DMS modification and one base was enhanced in the modification. The rRNA regions searched by nine primers and the detected bases are plotted on the 23S rRNA crystal structure of Deinococcus radiodurans (Harms et al. 2001; Protein Data Bank entry code 1NKW [PDB] ) as shown in Fig. 3. Figure 3a indicates that the searching regions in this study cover a wide range. Figure 3b indicates that many of the protected bases (red balls) are focused on a limited area. Interestingly A2451 is included among the protected bases in the area, which is thought to be directly involved in peptidyl transferase activity. From these observations, it is concluded that RMF binds at or near the PTase centre because the protected bases are focused on this area. The green and yellow tubes show the ribosomal proteins L2 and L13, respectively, and an intensively protected area exists between L2 and L13. These proteins were crosslinked with RMF in the previous studies (Yoshida et al. 2002). This fact shows that the results in this study are consistent with those from the protein-protein crosslinking study. The protected base (red ball) except the intensive area and the enhanced base (blue ball) are C2394 and A2432 at the upper left, respectively. These bases might be modified by effects of conformational changes.



View larger version (65K):
[in this window]
[in a new window]
  		Figure 2  Autoradiographs of primer extension analysis. Four lanes on the left, U, G, C and A, are dideoxy sequencing lanes. The next two are lanes of dimethyl sulphate (DMS) treatment with and without RMF, and the remaining two lanes are of non-DMS treatment with and without the RMF. The nucleotides that the DMS modification protected and enhanced are pointed out by black and white triangles, respectively. Stop signals at uridine bases are marked with asterisks, which are unlikely to reflect DMS-modifications. (a) A part of the region searched by using Pr-8 (Table 1). (b) By using Pr-6. (c) By using Pr-2. (d) By using Pr-1. (e) By using Pr-3. (f) By using Pr-4.

 


View larger version (40K):
[in this window]
[in a new window]
  		Figure 3  Detected bases on 23S rRNA tertiary structure of Deinococcus radiodurans. These figures show the views from the interface side of ribosomal large subunits. (a) Searched bases by nine primers are shown by green balls (see Table 1). (b) The bases protected from the DMS modification are shown by red balls, and the enhanced bases are shown by blue balls. A2451 (A2430 D. radiodurans numbering) is highlighted, which is thought to be directly involved in peptidyl transferase activity. The green and yellow tubes show the ribosomal proteins L2 and L13, respectively.

 

   Discussion
Top
 Abstract
 Introduction
 Results
 Discussion
Experimental procedures
 References
 
In order to investigate the functions of RMF, the binding sites on the 100S ribosome were examined by chemical probing using DMS. As a result, the nine protected bases and one enhanced base were found as shown in Fig. 2. When these bases were plotted on the 23S rRNA crystal structure, many of the protected bases were focused on the area around the PTase centre. This result indicates that RMF binds to a region at or near the PTase centre, which is consistent with the results of the crosslinking studies. Figure 4a is a predicted secondary structure of RMF by GTOP (Ito et al. 1997; http://spock.genes.nig.ac.jp/~genome/gtop.html). According to this prediction, RMF has two {alpha} helices at the N and C termini and a coil fastening the helices. We believe RMF has an elongated and simple structure, which can also be explained by instability of free-RMF in the cytoplasm. If this prediction is correct, the structural size of RMF will be able to cover the area identified as the binding sites. Homologous proteins of RMF have been identified in some gram-negative bacteria by using database-searching techniques as shown in Fig. 4b. There is considerable homology between them, indicating that their RMFs have the same binding sites as those of E. coli. Besides the above bacteria, the dimer of ribosomes has been experimentally found in Serratia marcescens and Proteus mirabilis (unpublished data). Most of them are enterobacteria, while P. aeruginosa and P. syringae are soil bacteria and their RMFs have an elongated C terminus. The relationship between the activity of RMF and the lifestyle of these bacteria is interesting. Some of the above bacteria are pathogenic bacteria. The RMF may become a candidate for targets for the design of new drugs to combat pathogenic bacteria.



View larger version (48K):
[in this window]
[in a new window]
  		Figure 4  Predicted secondary structure of RMF and comparison of RMF homologues. (a) The secondary structure of E. coli RMF is predicted by GTOP. The part except alpha-helix and beta-strand are represented as a coil. (b) Alignment of amino acid sequences of RMF from E. coli W3110 and O157, Shigella flexneri, Salmonella typhimurium LT2 and Ty2, Yersinia pestis, Vibrio cholerae, Vibrio parahaemolyticus, Pseudomonas aeruginosa, Pseudomonas syringae and Shewanella oneidensis.

 
The detected bases are far from each other on the secondary structure of 23S rRNA as shown in Fig. 5. They, however, stand close to each other on the tertiary structure as shown in Fig. 3. In Fig. 5, red and blue arrows show the bases that interact with the aminoacyl (A) site for aminoacyl-tRNA and the peptidyl (P) site for peptidyl-tRNA, respectively. This figure indicates that the protected bases by the RMF binding exist both near the A- and P-site. Interestingly, the RMF interacts with A2451. It is reported that A2451 plays an important role in PTase (Schmeing et al. 2002), and it is completely conserved among the three phylogenetic domains archaebacteria, eubacteria, and eukaryotes (Cannone et al. 2002). G2061 also exists near the PTase centre in the tertiary structure (Nissen et al. 2000), and its neighbouring bases, A2058, A2059 and A2062 are detected as the protected bases. It is reported that this region around G2061 is an entrance for the peptide exit tunnel, and these bases are highly conserved (Hansen et al. 2003). The bases near A1918 are far from the PTase centre on the secondary structure, but they are at the 30S-50S interface side of 23S rRNA over the PTase centre region on the tertiary structure and interact with the peptidyl tRNA as shown in Fig. 5a (Wilson et al. 2002). The A-site is the binding position of aminoacyl tRNA during the translational elongation step, and the P-site is of initiator aminoacyl tRNA during the translational initiation step and of peptidyl tRNA during the elongation step. According to these facts, it is assumed that RMF inhibits tRNA binding on to the ribosome. The translational activity of 100S ribosomes was examined in 1991, elucidating that RMF inhibits the binding of fMet-tRNA and Phe-tRNA in vitro (Wada et al. 1995). It is thought that fMet-tRNA binds to the P-site and that Phe-tRNA binds to the A-site and/or P-site in the experimental conditions. These data show that RMF occupies the P-site or both sites. The results of chemical probing in this study give meaningful interpretation to the fact that 100S ribosomes have no translational activity.



View larger version (31K):
[in this window]
[in a new window]
  		Figure 5  Detected bases on 23S rRNA secondary structure of E. coli. This figure was written by modifying data downloaded from http://www.rna.lcmb.utexas.edu. The bases protected from the DMS modification are shown by red balls, and the enhanced bases are shown by blue balls. Uridines detected in Figure 2 are marked with asterisks. The bases interacting with A- and P-site tRNAs, chloramphenicol, and erythromycin are also shown by red and blue arrows, and pink and green triangles, respectively. (a) In the vicinity of A1900–A1970. (b) In the vicinity of C2050–C2610.

 
It is known that some antibiotics inhibit the translational activity by binding near the PTase centre (Schlünzen et al. 2001). Chloramphenicol and erythromycin are the representative antibiotics among them. The bases that interacted with these antibiotics are shown by pink and green triangles in Fig. 5. These bases also overlap with the interaction sites of tRNA (A-site and P-site) and are close to the protected bases by RMF binding. It is reported that chloramphenicol and erythromycin predominantly bind to the hydrophobic crevices of the PTase centre and the entrance of the peptide exit tunnel, respectively (Hansen et al. 2003). The molecular weights of chloramphenicol and erythromycin are 323.13 and 733.94, respectively, which are smaller than that of RMF (MW: 6507). It is assumed that RMF binds to the ribosomes as though it covers the binding sites of these antibiotics, namely the PTase centre and the entrance of the peptide exit tunnel.

The DMS modification of one base was enhanced as shown in Fig. 3b. In general, it is explained that the enhancement of the chemical modification is caused by the conformational changes. On the basis of this interpretation, it is assumed that the modification of enhanced base detected in this study is caused by the conformational changes on the formation of 100S ribosomes. Also, it is assumed that the protected base except for the intensive area (C2394) escapes from DMS modification by the conformational changes. It is reported that RMF binds to the interface between the 50S and 30S subunits (Wada et al. 1990). However, in observations by electron microscopy it was elucidated that the 100S ribosome is formed by the dimerization of two 70S ribosomes mediated by face-to-face contacts between their constituent 30S subunits, namely 50S-30S-30S-50S (Yoshida et al. 2002). It is difficult to imagine how RMF with a molecular weight of only 6507 directly mediates the interaction between two 30S subunits. The intercalation of RMF in the interface may induce conformational changes in the 30S subunits, which causes two 70S ribosomes to dimerize. The protected and enhanced bases, except for the intensive area, may have relations with the conformational changes on the formation of the 100S ribosomes.

In conclusion, the binding site of RMF on E. coli 23S rRNA was identified by chemical probing. The detected bases in this study might include the results not only of direct protection by RMF but also of conformational changes. However, it is suggested that RMF binds near the PTase centre from the facts that many protected bases focus on the PTase centre and the results are consistent with crosslinking studies. These data provide a useful piece of information to illustrate the mechanism of translational inactivation of ribosomes by RMF. RMF binds to the ribosomes as though it covers the PTase centre and the entrance of the peptide exit tunnel or it occupies the P-site or both the A- and P-site. By the binding of RMF to the crucial sites, the ribosome is divested of the translational activity. The binding site of RMF is the same as some antibiotics. It is very surprising that the cell itself produces a protein that seems to inhibit protein synthesis in a similar manner to antibiotics and can moreover reversibly bind to and release from the ribosome in response to environmental conditions. The data in this study may provide information of the conformational changes on the formation of 100S ribosomes. When the 100S ribosome is formed, it is predicted that the 30S subunits undergo large conformational changes. The chemical probing method will be a useful tool to examine the conformational changes in detail. We are carrying out the structural analysis of the 100S ribosome by cryo electron microscopy and hoping that the mechanisms of dimerization are elucidated.


   Experimental procedures
Top
 Abstract
 Introduction
 Results
 Discussion
 Experimental procedures
References
 
Preparations of ribosomes and His-tagged RMF

E. coli W3110 cells were grown in medium E (Vogel & Bonner 1956) containing 2% polypeptone at 37 °C with shaking at 100 cycles per min for 4 days. Preparation of crude ribosomes (CR) from the cells was carried out essentially according to the method of Noll et al. (1973), with slight modifications as described by Horie et al. (1981). The CR, including the 100S ribosomes, was suspended in association buffer (100 mM CH3COONH4, 15 mM (CH3COO)2Mg, and 20 mM Tris-HCl at pH 7.6). High-salt washed ribosomes (HSR) were prepared by a centrifugation of CR in high-salt buffer (1 M CH3COONH4, 15 mM (CH3COO)2Mg, and 20 mM Tris-HCl at pH 7.6). The HSR includes mostly 70S ribosomes and does not include 100S ribosomes, because the RMF is released by this treatment and the 100S ribosome dissociates to two 70S ribosomes. The HSR prepared from the cells in the stationary phase has a higher efficiency of 100S ribosome formation in vitro using His-tagged RMF than that in the exponential phase. The CR and HSR were dialysed against DMS reaction buffer (100 mM KCl, 15 mM MgCl2, 50 mM K-cacodylate at pH 7.2) for use of DMS chemical probing.

The expression vector pQE-9 (QIAGEN), which contains a 6 x His-tag at the N-terminus, was used to express His-tagged RMF. The ligated vector [pQE-9(rmf-lacZ)] was transformed into M15 strains. M15 cells carrying pQE-9(rmf-lacZ) were grown in 2 x YT medium containing 25 µg/ml kanamycin and 100 µg/ml ampicillin at 37 °C with shaking at 100 cycles per min. Expression of His-tagged RMF was induced by adding IPTG [isopropyl-ß-D(-)-thiogalactopyranoside, Wako] to a final concentration of 0.03 mM at OD660nm 0.6. His-tagged RMF was purified by a column filled with 1 ml nickel-nitrilotriacetic acid-agarose (Ni-NTA, HiTrap column from Amersham Pharmacia Biotech.) and then was dialysed against DMS reaction buffer for use in DMS chemical probing.

Sucrose density gradient centrifugation

Ribosomes were subjected to centrifugation on 5–20% linear sucrose density gradients in DMS reaction buffer. After centrifugation in a SW40Ti rotor (Beckman) at 40 000 r.p.m. (285 000 g) for 80 min at 4 °C, ribosome profiles were observed at 260 nm by a UV-180 spectrometer (Shimazu) using a flow cell.

Chemical modification and primer extension

The 100S ribosomes formed in vitro were prepared by adding the His-tagged RMF at a 10-fold molar ratio to HSR, and by incubating at 37 °C for 30 min. The CR was employed as the native 100S ribosomes formed in vivo. The sample volumes of the ribosomal solutions (OD260nm: 0.5) were adjusted to 50 µl with DMS reaction buffer before chemical modification. Chemical modification was started by addition of 1 µL DMS (1 : 4 dilution in ethanol), and then incubated 37 °C for 10 min. RNA extraction, primer extension and gel electrophoresis were performed as described by Moazed and Noller (Moazed & Noller 1986). Nine primers were used in primer extension and they are listed in Table 1. Among them Pr-5 did not work in the extension reaction.


   Acknowledgements
 
We thank an anonymous reviewer for critical reading of the manuscript and helpful suggestions. This work was supported by Grants-in-Aid for Scientific Research on Priority Areas ‘Spatiotemporal Network of RNA Information Flow’ from the Ministry of Education, Culture, Sports, Science and Technology of Japan.


   Footnotes
 
Communicated by: Yoshikazu Nakamura

* Correspondence: E-mail: yhide{at}art.osaka-med.ac.jp


   References
Top
 Abstract
 Introduction
 Results
 Discussion
 Experimental procedures
 References
 
Cannone, J.J., Subramanian, S., Schnare, M.N., et al. (2002) The comparative RNA web (CRW) site: an online database of comparative sequence and structure information for ribosomal intron, and other RNA’s: Correction. BMC Bioinformat. 3, 2–31.[CrossRef][Medline]

Hansen, J.L., Moore, P.B. & Steitz, T.A. (2003) Structures of five antibiotics bound at the peptidyl transferase center of the large ribosomal subunit. J. Mol. Biol. 330, 1061–1075.[CrossRef][Medline]

Harms, J., Schluenzen, F., Zarivach, R., et al. (2001) High resolution structure of the large ribosomal subunit from a methophilic eubacterium. Cell 107, 679–688.[CrossRef][Medline]

Hinnebusch, A.G. (1994) Translational control of GCN4: an in vivo barometer of initiation-factor activity. Trends Biochem. Sci. 19, 409–414.[CrossRef][Medline]

Horie, K., Wada, A. & Fukutome, H. (1981) Conformational studies of Escherichia coli ribosomes with the use of acridine orange as a probe. J. Biochem. 90, 449–461.[Abstract/Free Full Text]

Ito, M., Matsuo, Y. & Nishikawa, K. (1997) Prediction of protein secondary structure using the 3D-1D compatibility algorithm. Comput. Appl. Biosci. 13, 415–424.[Abstract/Free Full Text]

Izutsu, K., Wada, A. & Wada, C. (2001a) Expression of ribosome modulation factor (RMF) in Escherichia coli requires ppGpp. Genes Cells 6, 665–676.[Abstract]

Izutsu, K., Wada, C., Komine, Y., et al. (2001b) Escherichia coli ribosome-associated protein SRA, whose copy number increases during stationary phase. J. Bacteriol. 183, 2765–2773.[Abstract/Free Full Text]

Maki, Y., Yoshida, H. & Wada, A. (2000) Two proteins, YfiA and YhbH, associated with resting ribosomes in stationary phase Escherichia coli. Genes Cells 5, 965–974.[Abstract]

Mizuta, K. & Warner, J.R. (1994) Continued functioning of the secretory pathway is essential for ribosome synthesis. Mol. Cell. Biol. 14, 2493–2502.[Abstract/Free Full Text]

Moazed, D. & Noller, H.F. (1986) Transfer RNA shields specific nucleotides in 16S ribosomal RNA from attack by chemical probes. Cell 47, 985–994.[CrossRef][Medline]

Nissen, P., Hansen, J., Ban, N., Moore, P.B. & Steitz, T.A. (2000) The structural basis of ribosome activity in peptide bond synthesis. Science 289, 920–930.[Abstract/Free Full Text]

Noll, M., Hapke, B., Schreier, M.H.K. & Noll, H. (1973) Structural dynamics of bacterial ribosomes. 1.Characterization of vacant couples and their relation to complexed ribosomes. J. Mol. Biol. 75, 281–294.[CrossRef][Medline]

Schlünzen, F., Zarivach, R., Harms, J., et al. (2001) Structural basis for the interaction of antibiotics with the peptidyl transferase centre in eubacteria. Nature 413, 814–821.[CrossRef][Medline]

Schmeing, T.M., Seila, A.C., Hansen, J.L., et al. (2002) A pre-translocational intermediate in protein synthesis observed in crystals of enzymatically active 50S subunits. Nature Struct. Biol. 9, 225–230.[Medline]

Vogel, H.J. & Bonner, D.M. (1956) Acetylornithinase of Escherichia coli: partial purification and some properties. J. Biol. Chem. 218, 97–106.[Free Full Text]

Wada, A. (1998) Growth phase coupled modulation of Escherichia coli ribosomes. Genes Cells 3, 203–208.[Abstract]

Wada, A., Igarashi, K., Yoshimura, S., Aimoto, S. & Ishihama, A. (1995) Ribosome modulation factor: Stationary growth phase-specific inhibitor of ribosome functions from Escherichia coli. Biochem. Biophys. Res. Commun. 214, 410–417.[CrossRef][Medline]

Wada, A., Yamazaki, Y., Fujita, N. & Ishihama, A. (1990) Structure and probable genetic location of a ‘ribosome modulation factor’ associated with 100S ribosomes in stationary-phase Escherichia coli cells. Proc. Natl. Acad. Sci. USA 87, 2657–2661.[Abstract/Free Full Text]

Wilson, D.N., Blaha, G., Connell, S.R., et al. (2002) Protein synthesis at atomic resolution: Mechanistics of translation in the light of highly resolved structures for the ribosome. Curr. Protein Pept. Sci. 3, 1–53.[Medline]

Yamagishi, M., Matsushima, H., Wada, A., Sakagami, M., Fujita, N. & Ishihama, A. (1993) Regulation of the Escherichia coli rmf gene encoding the ribosome modulation factor: growth phase- and growth rate-dependent control. EMBO J. 12, 625–630.[Medline]

Yoshida, H., Maki, Y., Kato, H., et al. (2002) The Ribosome Modulation Factor (RMF) Binding Site on the 100S Ribosome of Escherichia coli. J. Biochem. 132, 983–989.[Abstract/Free Full Text]

Received: 9 October 2003
Accepted: 15 January 2004




This article has been cited by other articles:


Home page
 J BiochemHome page
M. Ueta, R. L. Ohniwa, H. Yoshida, Y. Maki, C. Wada, and A. Wada
Role of HPF (Hibernation Promoting Factor) in Translational Activity in Escherichia coli
J. Biochem., March 1, 2008; 143(3): 425 - 433.
[Abstract] [Full Text] [PDF]

Home page
 MicrobiologyHome page
W. M. El-Sharoud and G. W. Niven
The influence of ribosome modulation factor on the survival of stationary-phase Escherichia coli during acid stress
Microbiology, January 1, 2007; 153(1): 247 - 253.
[Abstract] [Full Text] [PDF]

Home page
 GENES CELLSHome page
M. Ueta, H. Yoshida, C. Wada, T. Baba, H. Mori, and A. Wada
Ribosome binding proteins YhbH and YfiA have opposite functions during 100S formation in the stationary phase of Escherichia coli
Genes Cells, December 1, 2005; 10(12): 1103 - 1112.
[Abstract] [Full Text] [PDF]

Home page
 J. Bacteriol.Home page
P. Salunkhe, T. Topfer, J. Buer, and B. Tummler
Genome-Wide Transcriptional Profiling of the Steady-State Response of Pseudomonas aeruginosa to Hydrogen Peroxide
J. Bacteriol., April 15, 2005; 187(8): 2565 - 2572.
[Abstract] [Full Text] [PDF]

Home page
 J. Bacteriol.Home page
T. Aiso, H. Yoshida, A. Wada, and R. Ohki
Modulation of mRNA Stability Participates in Stationary-Phase-Specific Expression of Ribosome Modulation Factor
J. Bacteriol., March 15, 2005; 187(6): 1951 - 1958.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Yoshida, H.
Right arrow Articles by Wada, A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Yoshida, H.
Right arrow Articles by Wada, A.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE ADVANCED SEARCH TABLE OF CONTENTS