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Ribosomal Protection Proteins and Their Mechanism ofTetracyclineResistance
Open accessAbstract
Ribosomal protection represents an important tactic for promoting tetracycline resistance in both gram-positive and -negative species. Tet(O) and Tet(M) are the best studied of these determinants and were originally isolated from Campylobacter jejuni and Streptococcus spp., respectively, although both are widely distributed (10). These are the only two ribosomal protection proteins (RPPs) that have been studied in detail, and therefore, they have been dealt with extensively in this review. It is assumed, however, that the other members of this class of RPPs [Tet(S), Tet(T), Tet(Q), TetB(P), Tet(W), and OtrA] function through similar mechanisms. The distribution of these determinants in the eubacteria has been extensively reviewed by Chopra and Roberts (10) and more recent information can also be found at http://faculty.washington.edu/marilynr/. Although this review focuses primarily on RPPs, it should be noted that a great variety of tetracycline resistance mechanisms exist (for a review, see reference 10). These determinants include (i) the efflux-based mechanisms found in gram-positive and gram-negative bacteria (10), (ii) the enzymatic degradation of tetracyclines found in Bacteroides (46), (iii) the rRNA mutations found in Propionibacterium acnes and Helicobacter pylori (19, 40, 55), and (iv) a host of undetermined mechanisms which bear little resemblance to the well-documented determinants mentioned above (10). In this review, we will survey recent advances in the study of the ribosome, tetracycline, and the RPPs that further the understanding of RPP activity. Earlier work dealing with Tet(M) and Tet(O) as well as the other RPPs has been reviewed previously (51, 52).
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TL;DR
Ribosomal protection proteins (RPPs) such as Tet(O) and Tet(M) confer tetracycline resistance by binding the ribosome in a GTP-dependent manner and releasing tetracycline from the primary 30S binding site, restoring A-site tRNA accommodation. The review integrates biochemical and structural data into an allosteric model in which Tet(O) contacts h34 and perturbs the decoding center.
Key findings
- Tetracycline inhibits elongation by blocking aminoacyl-tRNA accommodation into the ribosomal A site; the high-affinity 30S binding site has a \(K_d\) of 1 to 20 µM, while crystal structures show tetracycline bound to either two or six sites on the 30S subunit. The primary/Tet-1 site near h34 and h31 is the likely inhibitory site.
- Tet(O) and Tet(M) are soluble cytoplasmic RPPs of ~72 kDa with 75% sequence similarity; they dislodge tetracycline from the ribosome and increase the apparent \(K_d\) of tetracycline binding from 5 to 30 µM, and Tet(M) protects A-site tRNA binding from tetracycline inhibition.
- RPPs bind the elongation factor site on the ribosome. A 16-Å cryo-EM reconstruction shows Tet(O) at the subunit interface, contacting mostly rRNA; its domain IV contacts h18/34 of 16S rRNA, unlike EF-G domain IV, which contacts H69 of 23S rRNA. DMS probing shows Tet(O) protects C1214 and enhances A1408.
- Proposed mechanism: Tet(O) binds a tetracycline-blocked ribosome with an open A site, triggers allosteric release of tetracycline via disturbance of h34, hydrolyzes GTP, and dissociates, returning the ribosome to the elongation cycle.
Methods
- Minireview synthesizing published biochemical, chemical probing, cryo-EM, and crystal structure studies; no new experimental data are reported.
- Focuses on Tet(O) and Tet(M), the only two RPPs studied in detail, and assumes other RPPs [Tet(S), Tet(T), Tet(Q), TetB(P), Tet(W), and OtrA] function through similar mechanisms.
- Main analysis integrates ribosome-tetracycline structures and RPP binding data into a model of tetracycline action and RPP-mediated tetracycline release.
Limitations
- Stated: only Tet(O) and Tet(M) have been studied in detail; other RPPs are assumed to function similarly.
- Stated: it remains unanswered whether RPPs actively prevent tetracycline rebinding after release; also, there is a discrepancy about GTP hydrolysis, as a nonhydrolyzable GTP analogue was active with Tet(O) but only partially active with Tet(M).
- Evident from the text: several mechanistic steps, including a tetracycline-induced conformational change and the role of h44 rearrangement, are proposed but not conclusively established.
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In the paper
“(iii) The crystal structures show that the face of tetracycline that interacts with the rRNA in the primary (and Tet-1) site is also the face where modifications result in a loss of biological activity (4, 39).”
What the sources say
- Not checked yetcites [4] 4. Brodersen, D. E., W. M. Clemons, A. P. Carter, R. J. Morgan-Warren, B. T. Wimberly, and V. Ramakrishnan. 2000. The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit. Cell 103: 1143-1154. source
- Not checked yetcites [39] 39. Rogalski, W. 1985. Chemical modification of the tetracyclines, p. 179-316. In J. H. Boothe and J. J. Hlavka (ed.), The tetracyclines (handbook of experimental pharmacology), vol. 78. Springer-Verlag, Berlin, Germany. source
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In the paper
“Nevertheless, the RPPs may be evolutionarily derived from the elongation factors, such that they lost their original function and have been adapted to function in tetracycline resistance.”
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In the paper
“This is significant, as two independent 30S subunit-tetracycline crystal structures show tetracycline bound to either two (4) or six sites (37) on the 30S ribosomal subunit (Fig. 2).”
What the sources say
- Not checked yetcites [4] 4. Brodersen, D. E., W. M. Clemons, A. P. Carter, R. J. Morgan-Warren, B. T. Wimberly, and V. Ramakrishnan. 2000. The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit. Cell 103: 1143-1154. source
- Not checked yetcites [37] 37. Pioletti, M., F. Schlunzen, J. Harms, R. Zarivach, M. Gluhmann, H. Avila, A. Bashan, H. Bartels, T. Auerbach, C. Jacobi, T. Hartsch, A. Yonath, and F. Franceschi. 2001. Crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and IF3. EMBO J. 20: 1829-1839. source
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In the paper
“The secondary and Tet-5 tetracycline-binding sites (Fig. 2) are also likely candidates for the inhibitory binding site (4, 37).”
What the sources say
- Not checked yetcites [4] 4. Brodersen, D. E., W. M. Clemons, A. P. Carter, R. J. Morgan-Warren, B. T. Wimberly, and V. Ramakrishnan. 2000. The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit. Cell 103: 1143-1154. source
- Not checked yetcites [37] 37. Pioletti, M., F. Schlunzen, J. Harms, R. Zarivach, M. Gluhmann, H. Avila, A. Bashan, H. Bartels, T. Auerbach, C. Jacobi, T. Hartsch, A. Yonath, and F. Franceschi. 2001. Crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and IF3. EMBO J. 20: 1829-1839. source
hedgedconditional- Asserted: passes — The authors put it forward as true.
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In the paper
“(ii) In the primary (and Tet-1) site, several tetracycline-RNA interactions are mediated through a magnesium ion which is known to be important for tetracycline binding (4, 57).”
What the sources say
- Not checked yetcites [4] 4. Brodersen, D. E., W. M. Clemons, A. P. Carter, R. J. Morgan-Warren, B. T. Wimberly, and V. Ramakrishnan. 2000. The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit. Cell 103: 1143-1154. source
- Not checked yetcites [57] 57. White, J. P., and C. R. Cantor. 1971. Role of magnesium in the binding of tetracycline to Escherichia coli ribosomes. J. Mol. Biol. 58: 397-400. source
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In the paper
“In this position, tetracycline cannot directly interfere with tRNA binding, but rather, it could exert its inhibitory effect by interfering with the transition between the open and closed states of the 30S ribosomal subunit, which is important for the decoding reaction (34).”
What the sources say
- Not checked yetcites [34] 34. Ogle, J. M., F. V. Murphy, M. J. Tarry, and V. Ramakrishnan. 2002. Selection of tRNA by the ribosome requires a transition from an open to a closed form. Cell 111: 721-732. source
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In the paper
“however, Burdett (5, 6) showed that Tet(M) cannot substitute for the elongation factors in vivo or in vitro.”
What the sources say
- Not checked yetcites [5] 5. Burdett, V. 1991. Purification and characterization of Tet(M), a protein that renders ribosomes resistant to tetracycline. J. Biol. Chem. 266: 2872-2877. source
- Not checked yetcites [6] 6. Burdett, V. 1996. Tet(M)-promoted release of tetracycline from ribosomes is GTP dependent. J. Bacteriol. 178: 3246-3251. source
plain- Asserted: passes — The authors put it forward as true.
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In the paper
“In this respect, the Tet-1 and primary site in the Pioletti and Brodersen structures, respectively (Fig. 2), are located in the ribosomal A site, where tetracycline is bound by the irregular minor groove of helix 34 (h34) and the loop of helix 31 (h31) in the 16S rRNA (4, 37).”
What the sources say
- Not checked yetcites [4] 4. Brodersen, D. E., W. M. Clemons, A. P. Carter, R. J. Morgan-Warren, B. T. Wimberly, and V. Ramakrishnan. 2000. The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit. Cell 103: 1143-1154. source
- Not checked yetcites [37] 37. Pioletti, M., F. Schlunzen, J. Harms, R. Zarivach, M. Gluhmann, H. Avila, A. Bashan, H. Bartels, T. Auerbach, C. Jacobi, T. Hartsch, A. Yonath, and F. Franceschi. 2001. Crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and IF3. EMBO J. 20: 1829-1839. source
plain- Asserted: passes — The authors put it forward as true.
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In the paper
“However, Tet(O), like the other RPPs, seems to have originated in the natural producer of oxytetracycline, Streptomyces rimosus , which harbors otrA , an RPP determinant (16, 44, 49).”
What the sources say
- Not checked yetcites [16] 16. Doyle, D., K. J. McDowall, M. J. Butler, and I. S. Hunter. 1991. Characterization of an oxytetracycline-resistance gene, otrA, of Streptomyces rimosus . Mol. Microbiol. 5: 2923-2933. source
- Not checked yetcites [44] 44. Sougakoff, W., B. Papadopoulou, P. Nordmann, and P. Courvalin. 1987. Nucleotide sequence and distribution of gene tetO encoding tetracycline resistance in Campylobacter coli . FEMS Microbiol. Lett. 44: 153-159. source
- Not checked yetcites [49] 49. Taylor, D. E. 1992. Antimicrobial resistance of Campylobacter jejuni and Campylobacter coli to tetracycline, chloramphenicol, and erythromycin, p. 74-86. In I. Nachaminkin (ed.), Campylobacter jejuni : current status and future trends. American Society for Microbiology, Washington, D.C.
hedgedcorrelational- Asserted: passes — The authors put it forward as true.
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In the paper
“Tet(O) and Tet(M) can dislodge tetracycline from the ribosome (6, 54) and, in so doing, increase the apparent dissociation constant ( Kd ) of tetracycline binding to the ribosome from 5 to 30 𝛍 M.”
What the sources say
- Not checked yetcites [6] 6. Burdett, V. 1996. Tet(M)-promoted release of tetracycline from ribosomes is GTP dependent. J. Bacteriol. 178: 3246-3251. source
- Not checked yetcites [54] 54. Trieber, C. A., N. Burkhardt, K. H. Nierhaus, and D. E. Taylor. 1998. Ribosomal protection from tetracycline mediated by Tet(O): Tet(O) interaction with ribosomes is GTP-dependent. Biol. Chem. 379: 847-855. source
plaindirect- Asserted: passes — The authors put it forward as true.
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- Probably not a substantial claim. 10
In the paper
“Accordingly, many of the RPP determinants are located on mobile genetic elements which may have facilitated their spread throughout the eubacteria via lateral gene transfer events (reviewed in reference 10).”
What the sources say
- Not checked yetcites [10] 10. Chopra, I., and M. Roberts. 2001. Tetracycline antibiotics: mode of action, applications, molecular biology, and epidemiology of bacterial resistance. Microbiol. Mol. Biol. Rev. 65: 232-260. source
hedgedconditional- Asserted: passes — The authors put it forward as true.
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In the paper
“Accordingly, the RPPs bind and hydrolyze GTP in a ribosome-dependent manner (5, 6, 53), and maintenance of this activity is important for in vivo activity (11, 23).”
What the sources say
- Not checked yetcites [5] 5. Burdett, V. 1991. Purification and characterization of Tet(M), a protein that renders ribosomes resistant to tetracycline. J. Biol. Chem. 266: 2872-2877. source
- Not checked yetcites [6] 6. Burdett, V. 1996. Tet(M)-promoted release of tetracycline from ribosomes is GTP dependent. J. Bacteriol. 178: 3246-3251. source
- Not checked yetcites [53] 53. Taylor, D. E., L. J. Jerome, J. Grewal, and N. Chang. 1995. Tet(O), a protein that mediates ribosomal protection to tetracycline, binds, and hydrolyses GTP. Can. J. Microbiol. 41: 965-970. source
plaindirect- Asserted: passes — The authors put it forward as true.
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It may state two separate claims as one.
- The authors state this as an assumption: The other members of the RPP class [Tet(S), Tet(T), Tet(Q), TetB(P), Tet(W), and OtrA] function through mechanisms similar to those of Tet(O) and Tet(M).
None of the sources cited for this claim have been checked yet.
In the paper
“Alternatively, the typical tetracyclines, which are the subject of RPP-mediated resistance, bind to the ribosome and inhibit the elongation phase of protein synthesis (8, 13).”
What the sources say
- Not checked yetcites [8] 8. Chopra, I. 1985. Mode of action of the tetracyclines and the nature of bacterial resistance to them, p. 317-392. In J. H. Boothe and J. J. Hlavka (ed.), The tetracyclines (handbook of experimental pharmacology), vol. 78. Springer-Verlag, Berlin, Germany. source
- Not checked yetcites [13] 13. Cundliffe, E. 1967. Antibiotics and polyribosomes. Chlortetracycline and polyribosomes of Bacillus megaterium . Mol. Pharmacol. 3: 401-411.
plaindirect- Asserted: passes — The authors put it forward as true.
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In the paper
“(vi) Tet(O), an RPP which confers resistance to tetracycline, chases tetracyclines from the primary binding site but not the secondary binding site (12).”
What the sources say
- Not checked yetcites [12] 12. Connell, S. R., C. A. Trieber, U. Stelzl, E. Einfeldt, D. E. Taylor, and K. H. Nierhaus. 2002. The tetracycline resistance protein, Tet(O), perturbs the conformation of the ribosomal decoding center. Mol. Microbiol. 45: 14631472. source
plaindirect- Asserted: passes — The authors put it forward as true.
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It may state two separate claims as one.
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In the paper
“(i) the primary (and Tet-1) site is the most highly occupied site in both structures showing tetracycline bound to the 30S subunit (4, 37) consistent with the idea that the high-affinity site is the inhibitory site (17, 47, 56).”
What the sources say
- Not checked yetcites [4] 4. Brodersen, D. E., W. M. Clemons, A. P. Carter, R. J. Morgan-Warren, B. T. Wimberly, and V. Ramakrishnan. 2000. The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit. Cell 103: 1143-1154. source
- Not checked yetcites [37] 37. Pioletti, M., F. Schlunzen, J. Harms, R. Zarivach, M. Gluhmann, H. Avila, A. Bashan, H. Bartels, T. Auerbach, C. Jacobi, T. Hartsch, A. Yonath, and F. Franceschi. 2001. Crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and IF3. EMBO J. 20: 1829-1839. source
- Not checked yetcites [17] 17. Epe, B., and P. Woolley. 1984. The binding of 6-demethylchlortetracycline to 70S, 50S and 30S ribosomal particles: a quantitative study by fluorescence anisotropy. EMBO J. 3: 121-126. source
hedgedcorrelational- Asserted: passes — The authors put it forward as true.
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It may state two separate claims as one.
In the paper
“RPPs, such as the well-studied Tet(O) and Tet(M) (75% sequence similarity), are soluble cytoplasmic proteins ( ∼ 72 kDa) which mediate tetracycline resistance (51).”
What the sources say
- Not checked yetcites [51] 51. Taylor, D. E., and A. Chau. 1996. Tetracycline resistance mediated by ribosomal protection. Antimicrob. Agents Chemother. 40: 1-5. source
plaindirect- Asserted: passes — The authors put it forward as true.
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- Load-bearing: unclear — It is unclear whether the paper’s conclusions depend on it.
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It may state two separate claims as one.
- The authors state this as an assumption: The other members of the RPP class [Tet(S), Tet(T), Tet(Q), TetB(P), Tet(W), and OtrA] function through mechanisms similar to those of Tet(O) and Tet(M).
None of the sources cited for this claim have been checked yet.
In the paper
“More precisely, they inhibit accommodation of aminoacyl-tRNA (aa-tRNA) into the ribosomal A site (Fig. 1, reactions a and b) and, therefore, prevent the addition of new amino acids to the growing polypeptide (22, 25, 29, 48).”
What the sources say
- Not checked yetcites [22] 22. Gordon, J. 1969. Hydrolysis of guanosine 5 ′ -triphosphate associated with binding of aminoacyl transfer ribonucleic acid to ribosomes. J. Biol. Chem. 244: 5680-5686. source
- Not checked yetcites [25] 25. Hierowski, M. 1965. Inhibition of protein synthesis by chlorotetracycline in the Escherichia coli in vitro system. Proc. Natl. Acad. Sci. USA 53: 594-599. source
- Not checked yetcites [29] 29. Lucas-Lenard, J., and A. L. Haenni. 1967. Requirement of guanosine 5 ′ -triphosphate for ribosomal binding of aminoacyl-sRNA. Biochemistry 59: 554-560.
- Not checked yetcites [48] 48. Suarez, G., and D. Nathans. 1965. Inhibition of aminoacyl tRNA binding to ribosomes by tetracycline. Biochem. Biophys. Res. Commun. 18: 743-750. source
plaindirect- Asserted: passes — The authors put it forward as true.
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In the paper
“Tet(O) was first cloned from a transferable plasmid pUA466 found in the food-borne pathogen C. jejuni (50).”
What the sources say
- Not checked yetcites [50] 50. Taylor, D. E. 1986. Plasmid-mediated tetracycline resistance in Campylobacter jejuni : expression in Escherichia coli and identification of homology with streptococcal class M determinant. J. Bacteriol. 165: 1037-1039. source
plain- Asserted: passes — The authors put it forward as true.
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Probably not a substantial claim. The paper’s conclusions may not depend on it.
In the paper
“RPPs, such as the well-studied Tet(O) and Tet(M) (75% sequence similarity), are soluble cytoplasmic proteins ( ∼ 72 kDa) which mediate tetracycline resistance (51).”
What the sources say
- Not checked yetcites [51] 51. Taylor, D. E., and A. Chau. 1996. Tetracycline resistance mediated by ribosomal protection. Antimicrob. Agents Chemother. 40: 1-5. source
plaincorrelational- Asserted: passes — The authors put it forward as true.
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Probably not a substantial claim. The paper’s conclusions may not depend on it.
None of the sources cited for this claim have been checked yet.
In the paper
“The RPPs display sequence similarity to the ribosomal elongation factors, EF-G and EF-Tu (41), and are grouped into the translation factor superfamily of GTPases (27).”
What the sources say
- Not checked yetcites [41] 41. Sanchez-Pescador, R., J. T. Brown, M. Roberts, and M. S. Urdea. 1988. Homology of the TetM with translational elongation factors: implications for potential modes of tetM -conferred tetracycline resistance. Nucleic Acids Res. 16: 1218. source
- Not checked yetcites [27] 27. Leipe, D. D., Y. I. Wolf, E. V. Koonin, and L. Aravind. 2002. Classification and evolution of P-loop GTPases and related ATPases. J. Mol. Biol. 317: 41-72. source
plaincorrelational- Asserted: passes — The authors put it forward as true.
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- Load-bearing: unclear — It is unclear whether the paper’s conclusions depend on it.
- Based: passes — The paper gives a reason why it holds.
It may state two separate claims as one.
- The authors state this as an assumption: The other members of the RPP class [Tet(S), Tet(T), Tet(Q), TetB(P), Tet(W), and OtrA] function through mechanisms similar to those of Tet(O) and Tet(M).
None of the sources cited for this claim have been checked yet.
In the paper
“These changes could be mediated by S12, which is in close proximity to the top of h44 (43, 58) and also appears to interact with Tet(O) (45).”
What the sources say
- Not checked yetcites [43] 43. Schluenzen, F., A. Tocilj, R. Zarivach, J. Harms, M. Gluehmann, D. Janell, A. Bashan, H. Bartels, I. Agmon, F. Franceschi, and A. Yonath. 2000. Structure of functionally activated small ribosomal subunit at 3.3 angstroms resolution. Cell 102: 615-623. source
- Not checked yetcites [58] 58. Wimberly, B. T., D. E. Brodersen, W. M. Clemons, Jr., R. J. Morgan-Warren, A. P. Carter, C. Vonrhein, T. Hartsch, and V. Ramakrishnan. 2000. Structure of the 30S ribosomal subunit. Nature 407: 327-339. source
- Not checked yetcites [45] 45. Spahn, C. M. T., G. Blaha, R. K. Agrawal, P. Penczek, R. A. Grassucci, C. A. Trieber, S. R. Connell, D. E. Taylor, K. H. Nierhaus, and J. Frank. 2001. Localization of the ribosomal protection protein Tet(O) on the ribosome and the mechanism of tetracycline resistance. Mol. Cell 7: 1037-1045. source
hedgedconditional- Asserted: passes — The authors put it forward as true.
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It may state two separate claims as one.
In the paper
“Spahn et al. (45) proposed that Tet(O) triggers the release of tetracycline through an allosteric mechanism.”
What the sources say
- Not checked yetcites [45] 45. Spahn, C. M. T., G. Blaha, R. K. Agrawal, P. Penczek, R. A. Grassucci, C. A. Trieber, S. R. Connell, D. E. Taylor, K. H. Nierhaus, and J. Frank. 2001. Localization of the ribosomal protection protein Tet(O) on the ribosome and the mechanism of tetracycline resistance. Mol. Cell 7: 1037-1045. source
hedgeddirect- Asserted: passes — The authors put it forward as true.
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Probably not a substantial claim.
None of the sources cited for this claim have been checked yet.
In the paper
“The protection of C1214 from DMS modification would indicate that this base is directly shielded by Tet(O), and this conclusion is supported by the fact that the Tet(O) binding site observed by cryo-EM approaches C1214 (Fig. 4B).”
What the sources say
- Not checked yetcites [12] 12. Connell, S. R., C. A. Trieber, U. Stelzl, E. Einfeldt, D. E. Taylor, and K. H. Nierhaus. 2002. The tetracycline resistance protein, Tet(O), perturbs the conformation of the ribosomal decoding center. Mol. Microbiol. 45: 14631472. source
- Not checked yetcites [45] 45. Spahn, C. M. T., G. Blaha, R. K. Agrawal, P. Penczek, R. A. Grassucci, C. A. Trieber, S. R. Connell, D. E. Taylor, K. H. Nierhaus, and J. Frank. 2001. Localization of the ribosomal protection protein Tet(O) on the ribosome and the mechanism of tetracycline resistance. Mol. Cell 7: 1037-1045. source
hedgeddirect- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: passes — The paper’s conclusions depend on it.
- Based: passes — The paper gives a reason why it holds.
It may state two separate claims as one.
A conclusion the paper draws from its own results — it cites no other source for it, so there is none to check. The quote shows it is in the paper.
In the paper
“This, however, probably does not reflect a functional difference, but instead, it may result from the different methods used in each laboratory.”
hedgedconditional- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: fails — The paper’s conclusions may not depend on it.
- Based: passes — The paper gives a reason why it holds.
Probably not a substantial claim. The paper’s conclusions may not depend on it.
In the paper
“In the presence of tetracycline, the ribosome is blocked with an open A site and this could provide a kinetic window for Tet(O) to act, thus distinguishing the tetracycline-blocked ribosome from a translating ribosome (11).”
What the sources say
- Not checked yetcites [11] 11. Connell, S. R., C. A. Trieber, E. Einfeldt, G. P. Dinos, D. E. Taylor, and K. Nierhaus. 2003. Mechanism of Tet(O)-mediated tetracycline resistance. EMBO J. 22: 945-953. source
hedgedconditional- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: passes — The paper’s conclusions depend on it.
- Based: passes — The paper gives a reason why it holds.
It may state two separate claims as one.
None of the sources cited for this claim have been checked yet.
In the paper
“In this case, these differences in domain IV may serve to distinguish Tet(O) and EF-G with respect to their activities; namely, domain IV of EF-G more intimately overlaps with the A site-bound tRNA, an idea that is consistent with the role of domain IV of EF-G in translocation. In contrast, the interaction of domain IV of Tet(O) and h34 of the 30S subunit is consistent with its role in tetracycl”
What the sources say
- Not checked yetcites [45] 45. Spahn, C. M. T., G. Blaha, R. K. Agrawal, P. Penczek, R. A. Grassucci, C. A. Trieber, S. R. Connell, D. E. Taylor, K. H. Nierhaus, and J. Frank. 2001. Localization of the ribosomal protection protein Tet(O) on the ribosome and the mechanism of tetracycline resistance. Mol. Cell 7: 1037-1045. source
- Not checked yetcites [4] 4. Brodersen, D. E., W. M. Clemons, A. P. Carter, R. J. Morgan-Warren, B. T. Wimberly, and V. Ramakrishnan. 2000. The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit. Cell 103: 1143-1154. source
- Not checked yetcites [37] 37. Pioletti, M., F. Schlunzen, J. Harms, R. Zarivach, M. Gluhmann, H. Avila, A. Bashan, H. Bartels, T. Auerbach, C. Jacobi, T. Hartsch, A. Yonath, and F. Franceschi. 2001. Crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and IF3. EMBO J. 20: 1829-1839. source
hedgedconditional- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: passes — The paper’s conclusions depend on it.
- Based: passes — The paper gives a reason why it holds.
It may state two separate claims as one.
In the paper
“Spahn et al. (45) hypothesize that, upon binding, Tet(O) contacts the base of h34, which in turn causes a disturbance in this helix, which is propagated to the primary tetracycline-binding site, releasing the drug.”
What the sources say
- Not checked yetcites [45] 45. Spahn, C. M. T., G. Blaha, R. K. Agrawal, P. Penczek, R. A. Grassucci, C. A. Trieber, S. R. Connell, D. E. Taylor, K. H. Nierhaus, and J. Frank. 2001. Localization of the ribosomal protection protein Tet(O) on the ribosome and the mechanism of tetracycline resistance. Mol. Cell 7: 1037-1045. source
hedgeddirect- Asserted: fails — The authors may not put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: passes — The paper’s conclusions depend on it.
- Based: passes — The paper gives a reason why it holds.
Probably not a substantial claim. The authors may not put it forward as true. It may state two separate claims as one.
In the paper
“In the cryo-EM study, Spahn et al. localize the sites of interaction between Tet(O) and the ribosome (Table 1), demonstrating that that the majority of the interactions are between Tet(O) and the rRNA (45). The only exception is a single interaction between domain III of Tet(O) and the ribosomal protein S12 (45).”
What the sources say
- Not checked yetcites [45] 45. Spahn, C. M. T., G. Blaha, R. K. Agrawal, P. Penczek, R. A. Grassucci, C. A. Trieber, S. R. Connell, D. E. Taylor, K. H. Nierhaus, and J. Frank. 2001. Localization of the ribosomal protection protein Tet(O) on the ribosome and the mechanism of tetracycline resistance. Mol. Cell 7: 1037-1045. source
boostedcorrelational- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: passes — The paper’s conclusions depend on it.
- Based: passes — The paper gives a reason why it holds.
It may state two separate claims as one.
None of the sources cited for this claim have been checked yet.
In the paper
“It should be noted, however, that a direct interaction between Tet(O) and tetracycline bound to the primary site cannot be absolutely discounted, although both chemical probing and cryo-EM suggest this is not the case (12, 45).”
What the sources say
- Not checked yetcites [12] 12. Connell, S. R., C. A. Trieber, U. Stelzl, E. Einfeldt, D. E. Taylor, and K. H. Nierhaus. 2002. The tetracycline resistance protein, Tet(O), perturbs the conformation of the ribosomal decoding center. Mol. Microbiol. 45: 14631472. source
- Not checked yetcites [45] 45. Spahn, C. M. T., G. Blaha, R. K. Agrawal, P. Penczek, R. A. Grassucci, C. A. Trieber, S. R. Connell, D. E. Taylor, K. H. Nierhaus, and J. Frank. 2001. Localization of the ribosomal protection protein Tet(O) on the ribosome and the mechanism of tetracycline resistance. Mol. Cell 7: 1037-1045. source
hedged- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: unclear — It is unclear whether the paper’s conclusions depend on it.
- Based: passes — The paper gives a reason why it holds.
It may state two separate claims as one.
A conclusion the paper draws from its own results — it cites no other source for it, so there is none to check. The quote shows it is in the paper.
In the paper
“Thus, it appears that Tet(O) and Tet(M) confer tetracycline resistance by releasing tetracycline from the ribosome and thereby freeing the ribosome from the inhibitory effects of the drug, such that aa-tRNA can bind to the A site and protein synthesis can continue.”
hedgeddirect- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: passes — The paper’s conclusions depend on it.
- Based: passes — The paper gives a reason why it holds.
It may state two separate claims as one.
In the paper
“These bases are distinct from the observed tetracycline binding sites and may indicate that tetracycline is promoting subtle structural rearrangement or fixing the ribosome in a conformation that is favorable for the establishment of the C1402-C1501 cross-link.”
What the sources say
- Not checked yetcites [32] 32. Noah, J. W., M. A. Dolan, P. Babin, and P. Wollenzien. 1999. Effects of tetracycline on the tertiary structure of ribosomal RNA in the Escherichia coli 30S ribosomal subunit. J. Biol. Chem. 274: 16576-16581. source
hedgedconditional- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: fails — The paper’s conclusions may not depend on it.
- Based: passes — The paper gives a reason why it holds.
Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.
A conclusion the paper draws from its own results — it cites no other source for it, so there is none to check. The quote shows it is in the paper.
In the paper
“Additionally, as h44 is a component of the decoding site, it may have a role in the inhibitory action of tetracycline.”
hedgedconditional- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: fails — The paper’s conclusions may not depend on it.
- Based: passes — The paper gives a reason why it holds.
Probably not a substantial claim. The paper’s conclusions may not depend on it. It may state two separate claims as one.
A conclusion the paper draws from its own results — it cites no other source for it, so there is none to check. The quote shows it is in the paper.
In the paper
“the change might simply move the ribosome into a configuration compatible with stable tetracycline binding.”
hedgedconditional- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: fails — The paper’s conclusions may not depend on it.
- Based: passes — The paper gives a reason why it holds.
Probably not a substantial claim. The paper’s conclusions may not depend on it.
A conclusion the paper draws from its own results — it cites no other source for it, so there is none to check. The quote shows it is in the paper.
In the paper
“it is possible that the binding of tetracycline to the ribosome (reaction f) is accompanied by a structural rearrangement.”
hedgedconditional- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: fails — The paper’s conclusions may not depend on it.
- Based: passes — The paper gives a reason why it holds.
Probably not a substantial claim. The paper’s conclusions may not depend on it.
In the paper
“Noah et al. (32) presented evidence that tetracycline affects h44.”
What the sources say
- Not checked yetcites [32] 32. Noah, J. W., M. A. Dolan, P. Babin, and P. Wollenzien. 1999. Effects of tetracycline on the tertiary structure of ribosomal RNA in the Escherichia coli 30S ribosomal subunit. J. Biol. Chem. 274: 16576-16581. source
plaindirect- Asserted: passes — The authors put it forward as true.
- Checkable: passes — It can be checked against evidence.
- Load-bearing: fails — The paper’s conclusions may not depend on it.
- Based: passes — The paper gives a reason why it holds.
Probably not a substantial claim. The paper’s conclusions may not depend on it.
The authors state this as an assumption.
The other members of the RPP class [Tet(S), Tet(T), Tet(Q), TetB(P), Tet(W), and OtrA] function through mechanisms similar to those of Tet(O) and Tet(M).
“It is assumed, however, that the other members of this class of RPPs [Tet(S), Tet(T), Tet(Q), TetB(P), Tet(W), and OtrA] function through similar mechanisms.”
The paper relies on this without support.
After being released, tetracycline is free to rebind the ribosome and again inhibit protein synthesis.
“after being released, tetracycline is free to rebind the ribosome and again inhibit protein synthesis.”
Claims other papers make about this one
Each is checked against this paper’s own text: does it say what the citing paper says it does?
In the citing paper
“Tet(M) was initially described in Streptococcus spp. and Tet(O) in Campylobacter jejuni, but they are now both widely distributed among different bacterial species, likely because they have been found in several plasmids and in broad-range conjugative transposons (51).”
From Mechanisms of Antibiotic Resistance (2016) · cites this paper as [51]
In the citing paper
“As TetO is not observed to directly interact with C1054 or A1408 (14), TetO was suggested to chase tetracycline from the ribosome indirectly via inducing local disturbances within h34 (9,14,15).”
From Structural basis for TetM-mediated tetracycline resistance (2012) · cites this paper as [9]
In the citing paper
“Moreover, the conformational changes were proposed to persist after TetO has dissociated from the ribosome, preventing rebinding of tetracycline as well as stimulating delivery of the ternary complex (9,16).”
From Structural basis for TetM-mediated tetracycline resistance (2012) · cites this paper as [9]
Citations
Cites
VMD: visual molecular dynamics
Humphrey, W., Dalke, A., & Schulten, K. (1996) · Journal of Molecular Graphics · doi:10.1016/0263-7855(96)00018-5 · Cited by 66,952
Chopra, I., & Roberts, M. (2001) · Microbiology and Molecular Biology Reviews · doi:10.1128/mmbr.65.2.232-260.2001 · Cited by 4,517
The Complete Atomic Structure of the Large Ribosomal Subunit at 2.4 Å Resolution
Ban, N., Nissen, P., Hansen, J., Moore, P. B., & Steitz, T. A. (2000) · Science · doi:10.1126/science.289.5481.905 · Cited by 3,327
Structure of the 30S ribosomal subunit
Wimberly, B. T., Brodersen, D. E., Clemons, W. M., Morgan-Warren, R. J., Carter, A. P., Vonrhein, C., Hartsch, T., & Ramakrishnan, V. (2000) · Nature · doi:10.1038/35030006 · Cited by 2,035
Classification and evolution of P-loop GTPases and related ATPases
Leipe, D. D., Wolf, Y. I., Koonin, E. V., & Aravind, L. (2002) · Journal of Molecular Biology · doi:10.1006/jmbi.2001.5378 · Cited by 1,148
Structure of Functionally Activated Small Ribosomal Subunit at 3.3 Å Resolution
Schluenzen, F., Tocilj, A., Zarivach, R., Harms, J., Gluehmann, M., Janell, D., Bashan, A., Bartels, H., Agmon, I., Franceschi, F., et al. (2000) · Cell · doi:10.1016/s0092-8674(00)00084-2 · Cited by 979
Brodersen, D. E., Clemons, W. M., Carter, A. P., Morgan-Warren, R. J., Wimberly, B. T., & Ramakrishnan, V. (2000) · Cell · doi:10.1016/s0092-8674(00)00216-6 · Cited by 950 · cited 10×
“(4) postulated that with tetracycline bound to the primary site, the ternary complex would be able to initiate decoding, such that the interaction between the codon and the anticodon of the EF-Tu-bound aa-tRNA would be unaffected by the presence of the drug (Fig.”
BackgroundInfluentialHigh Resolution Structure of the Large Ribosomal Subunit from a Mesophilic Eubacterium
Harms, J. M., Schluenzen, F., Zarivach, R., Bashan, A., Gat, S., Agmon, I., Bartels, H., Franceschi, F. J., & Yonath, A. (2001) · Cell · doi:10.1016/s0092-8674(01)00546-3 · Cited by 899
Selection of tRNA by the Ribosome Requires a Transition from an Open to a Closed Form
Ogle, J. M., Murphy, F. V., Tarry, M. J., & Ramakrishnan, V. (2002) · Cell · doi:10.1016/s0092-8674(02)01086-3 · Cited by 663
Crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and IF3
Pioletti, M., Schlünzen, F., Harms, J., Zarivach, R., Glühmann, M., Avila, H., Bashan, A., Bartels, H., Auerbach, T., Jacobi, C., et al. (2001) · The EMBO Journal · doi:10.1093/emboj/20.8.1829 · Cited by 527
Cited by
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Mechanisms of Antibiotic Resistance
Munita, J. M., & Arias, C. A. (2016) · Microbiology Spectrum · doi:10.1128/microbiolspec.vmbf-0016-2015 · Cited by 2,782
Multidrug Resistance in Bacteria
Nikaido, H. (2009) · Annual Review of Biochemistry · doi:10.1146/annurev.biochem.78.082907.145923 · Cited by 1,776
Coagulase-Negative Staphylococci
Becker, K., Heilmann, C., & Peters, G. (2014) · Clinical Microbiology Reviews · doi:10.1128/cmr.00109-13 · Cited by 1,742
Molecular Mechanisms of Antibacterial Multidrug Resistance
Alekshun, M. N., & Levy, S. B. (2007) · Cell · doi:10.1016/j.cell.2007.03.004 · Cited by 1,708
Ribosome-targeting antibiotics and mechanisms of bacterial resistance
Wilson, D. N. (2014) · Nature Reviews Microbiology · doi:10.1038/nrmicro3155 · Cited by 1,054
Metadata
- Authors
- Connell, S. R., Tracz, D. M., Nierhaus, K. H., & Taylor, D. E.
- Affiliations
- University of Alberta (CA)
- Max Planck Institute for Molecular Genetics (DE)
- Venue
- Antimicrobial Agents and Chemotherapy
- Publisher
- American Society for Microbiology
- Published
- 2003-12
- Volume
- 47
- Issue
- 12
- Pages
- 3675-3681
- ISSN
- 0066-4804 (ISSN-L)
- 1098-6596
- PMC
- PMC296194
- OpenAlex
- W2168967909
- Semantic Scholar
- b51e42f14821598e8ac23c8bbd012170a3de473f
- Cited by
- 449
- References
- 60
- Citations by year
- 2026: 22 · 2025: 35 · 2024: 20 · 2023: 18 · 2022: 19 · 2021: 28 · 2020: 20 · 2019: 29 · 2018: 31 · 2017: 28 · 2016: 23 · 2015: 21 · 2014: 23 · 2013: 17 · 2012: 17
- FWCI
- 0.83
- Type
- journal article
- Language
- en
- Open access
- green
- europepmc.org
- Locations
- Antimicrobial Agents and Chemotherapy (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Max Planck Institute for Plasma Physics (submittedVersion)
- Topics
- Health Sciences › Medicine › Microbiology (medical) › Antibiotic Resistance in Bacteria
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Keywords
- ribosomal protection proteins, tetracycline resistance, tetM, tetO, ribosome protection, Campylobacter jejuni, Streptococcus, rRNA mutations, OTRA, tetW
- MeSH terms
- Bacteria, Bacterial Proteins, Models, Molecular, Ribosomes, Tetracycline Resistance
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
A language model drew the claims from the paper on 2026-10-10, under version 3 of the claim definition (model deepseek-v4.1-flash, prompt extract-v3).
A second model labelled each claim on 2026-10-10 (model jev-1.13-20260917, definition revision gold-guideline-v0.2).
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
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