Ribosomal RNA

Nucleoside modifications in rRNA

Posttranscriptional modification in rRNA was recognized and reported in the early literature, but has received far less attention than for tRNA because of limited sequence studies carried out at the RNA level (which was historically a frequent means of discovery of new nucleosides in tRNA), coupled with considerably less understanding of modification structure-function relationships compared with tRNA. In compiling a comprehensive and reliable listing of modified nucleosides in rRNA, several problems arose that were common to all of the RNAs but were perhaps more prevalent for rRNAs. First, some early work predated an understanding of the structures of rRNAs and of methods to isolate them without cross-contamination from other rRNAs, or from tRNA. Second, methods of structural identification prior to about 1965 were inadequate in the event that structurally new or unexpected nucleosides were encountered, and some identifications based principally on chromatographic behavior must be regarded as inconclusive even though no outright assignment error is apparent. In addition, RNA hydrolysis using acid or base (rather than nucleases) must have inevitably led to some formation of degraded or rearranged products, and loss of labile substituents, which were not readily recognized as such. As a consequence of these problems, some of the occurrences listed in rRNA are tenuous (as indicated by appropriate Comments in certain files) and should be verified using rigorous experimental methods.

Because of the relatively greater extent to which modifications in E. coli rRNA have been studied, it is possible to compare earlier reports with more recent work (e.g., 1), and thereby exclude one-time or unusual assignments that have not been subsequently verified by independent methods. On this basis, the following reported nucleoside occurrences in E. coli rRNA are not included in the database: m22G [2], and D [3] and I [3] in 16S, and I [3], m4C [4] and m3U [4] in 23S. In spite of the problems outlined above, and because of the absence of any attempt prior to 1994 to compile a comprehensive listing of rRNA modifications, the view has been adopted that some unusual occurrences in rRNA should be included in the database, even though uncertainties remain.

Modifications in E. coli 16S and 23S rRNA

The sites and identities of posttranscriptional modifications in the E. coli rRNAs are the most extensively studied of any prokaryote. A summary of currently known modifications, which is probably complete, is given in the table below. Some earlier assignments proved to be incorrect (as discussed in the section above and in ref. 5), sometimes as a consequence of unavailability at that time of the corresponding gene sequences which led to numerous published errors. Leading references for the data presented in the table below can be found in refs. 1, 6 (16S) and 7,8,9,10 (23S). Data for E. coli 5S rRNA is absent in the table because this rRNA is unmodified (e.g., ref. 1), a characteristic of most 5 - 5.8S rRNAs.

Modifications in E. coli rRNA
16S
Location Modification
516 Y
527 m7G
966 m2G
967 m5C
1207 m2G
1402 m4Cm
1407 m5C
1498 m3U
1516 m2G
1518 m62A
1519 m62A
23S
Location Modification
745 m1G
746 Y
747 m5U (T)
955 Y
1618 m6A
1835 m2G
1911 Y
1915 m3Y
1917 Y
1939 m5U (T)
1962 m5C
2030 m6A
2069 m7G
2251 Gm
2445 m2G
2449 D
2457 Y
2498 Cm
2501 C* (a)
2503 m2A
2504 Y
2552 Um
2580 Y
2605 Y
(a) C* is a substoichiometric
modification [10].

References for the Ribosomal RNA section

  1. Kowalak, J.A., Pomerantz, S.C., Crain, P.F. and McCloskey, J.A. (1993) A novel method for the determination of post-transcriptional modification in RNA by mass spectrometry. Nucleic Acids Res, 21, pp. 4577-85. PubMed
  2. Starr, J.L. and Fefferman, R. (1964) The Occurrence of Methylated Bases in Ribosomal Ribonucleic Acid of Escherichia coli K12 W-6. J Biol Chem, 239, pp. 3457-61. PubMed
  3. Johnson, J.D. and Horowitz, J. (1971) Characterization of ribosomes and RNAs from Mycoplasma hominis. Biochim Biophys Acta, 247, pp. 262-79. PubMed
  4. Gehrke, C.W. and Kuo, K.C. (1989) Ribonucleoside analysis by reversed-phase high-performance liquid chromatography. J Chromatogr, 471, pp. 3-36. PubMed
  5. Limbach, P.A., Crain, P.F. and McCloskey, J.A. (1994) Summary: the modified nucleosides of RNA. Nucleic Acids Res, 22, pp. 2183-96. PubMed
  6. Bakin, A., Kowalak, J.A., McCloskey, J.A. and Ofengand, J. (1994) The single pseudouridine residue in Escherichia coli 16S RNA is located at position 516. Nucleic Acids Res, 22, pp. 3681-4. PubMed
  7. Kowalak, J.A., Bruenger, E. and McCloskey, J.A. (1995) Posttranscriptional modification of the central loop of domain V in Escherichia coli 23 S ribosomal RNA. J Biol Chem, 270, pp. 17758-64. PubMed
  8. Branlant, C., Krol, A., Machatt, M.A., Pouyet, J., Ebel, J.P., Edwards, K. and Kossel, H. (1981) Primary and secondary structures of Escherichia coli MRE 600 23S ribosomal RNA. Comparison with models of secondary structure for maize chloroplast 23S rRNA and for large portions of mouse and human 16S mitochondrial rRNAs. Nucleic Acids Res, 9, pp. 4303-24. PubMed
  9. Bakin, A. and Ofengand, J. (1993) Four newly located pseudouridylate residues in Escherichia coli 23S ribosomal RNA are all at the peptidyltransferase center: analysis by the application of a new sequencing technique. Biochemistry, 32, pp. 9754-62. PubMed
  10. Andersen, T.E., Porse, B.T. and Kirpekar, F. (2004) A novel partial modification at C2501 in Escherichia coli 23S ribosomal RNA. RNA, 10, pp. 907-13. PubMed