Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1995 Dec 5;92(25):11681–11685. doi: 10.1073/pnas.92.25.11681

The "DEAD box" protein DbpA interacts specifically with the peptidyltransferase center in 23S rRNA.

S M Nicol 1, F V Fuller-Pace 1
PMCID: PMC40466  PMID: 8524828

Abstract

The Escherichia coli DEAD (Asp-Glu-Ala-Asp) box protein DbpA is a putative RNA helicase and established RNA-dependent ATPase and is the only member of the DEAD box protein family for which a specific RNA substrate, bacterial 23S rRNA, has been identified. We have investigated the nature of this specificity in depth and have localized by deletion mutagenesis and PCR a single region of 93 bases (bases 2496-2588) in 23S rRNA that is both necessary and sufficient for complete activation of ATPase activity of DbpA. This target region forms part of the peptidyltransferase center and includes many bases involved in interaction with the 3' terminal adenosines of both A- and P-site tRNAs. Deletion of stem loops within the 93-base segment abolished ATPase activation. Similarly, point mutations that disrupt base pairing within stem structures ablated stimulation of ATPase activity. These data are consistent with roles for DbpA either in establishing and/or maintaining the correct three-dimensional structure of the peptidyltransferase center in 23S rRNA during ribosome assembly or in the peptidyltransferase reaction.

Full text

PDF
11681

Images in this article

11682Fig. 1
on p.11682
11683Fig. 2
on p.11683

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Brosius J., Ullrich A., Raker M. A., Gray A., Dull T. J., Gutell R. R., Noller H. F. Construction and fine mapping of recombinant plasmids containing the rrnB ribosomal RNA operon of E. coli. Plasmid. 1981 Jul;6(1):112–118. doi: 10.1016/0147-619x(81)90058-5. [DOI] [PubMed] [Google Scholar]
  2. Chang T. H., Arenas J., Abelson J. Identification of five putative yeast RNA helicase genes. Proc Natl Acad Sci U S A. 1990 Feb;87(4):1571–1575. doi: 10.1073/pnas.87.4.1571. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Dohme F., Nierhaus K. H. Total reconstitution and assembly of 50 S subunits from Escherichia coli Ribosomes in vitro. J Mol Biol. 1976 Nov 15;107(4):585–599. doi: 10.1016/s0022-2836(76)80085-x. [DOI] [PubMed] [Google Scholar]
  4. Fuller-Pace F. V., Nicol S. M., Reid A. D., Lane D. P. DbpA: a DEAD box protein specifically activated by 23s rRNA. EMBO J. 1993 Sep;12(9):3619–3626. doi: 10.1002/j.1460-2075.1993.tb06035.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Fuller-Pace F. V. RNA helicases: modulators of RNA structure. Trends Cell Biol. 1994 Aug;4(8):271–274. doi: 10.1016/0962-8924(94)90210-0. [DOI] [PubMed] [Google Scholar]
  6. Grifo J. A., Abramson R. D., Satler C. A., Merrick W. C. RNA-stimulated ATPase activity of eukaryotic initiation factors. J Biol Chem. 1984 Jul 10;259(13):8648–8654. [PubMed] [Google Scholar]
  7. Gururajan R., Mathews L., Longo F. J., Weeks D. L. An3 mRNA encodes an RNA helicase that colocalizes with nucleoli in Xenopus oocytes in a stage-specific manner. Proc Natl Acad Sci U S A. 1994 Mar 15;91(6):2056–2060. doi: 10.1073/pnas.91.6.2056. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Herold M., Nierhaus K. H. Incorporation of six additional proteins to complete the assembly map of the 50 S subunit from Escherichia coli ribosomes. J Biol Chem. 1987 Jun 25;262(18):8826–8833. [PubMed] [Google Scholar]
  9. Hirling H., Scheffner M., Restle T., Stahl H. RNA helicase activity associated with the human p68 protein. Nature. 1989 Jun 15;339(6225):562–564. doi: 10.1038/339562a0. [DOI] [PubMed] [Google Scholar]
  10. Iggo R. D., Jamieson D. J., MacNeill S. A., Southgate J., McPheat J., Lane D. P. p68 RNA helicase: identification of a nucleolar form and cloning of related genes containing a conserved intron in yeasts. Mol Cell Biol. 1991 Mar;11(3):1326–1333. doi: 10.1128/mcb.11.3.1326. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Iggo R. D., Lane D. P. Nuclear protein p68 is an RNA-dependent ATPase. EMBO J. 1989 Jun;8(6):1827–1831. doi: 10.1002/j.1460-2075.1989.tb03577.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Iggo R., Picksley S., Southgate J., McPheat J., Lane D. P. Identification of a putative RNA helicase in E.coli. Nucleic Acids Res. 1990 Sep 25;18(18):5413–5417. doi: 10.1093/nar/18.18.5413. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kalman M., Murphy H., Cashel M. rhlB, a new Escherichia coli K-12 gene with an RNA helicase-like protein sequence motif, one of at least five such possible genes in a prokaryote. New Biol. 1991 Sep;3(9):886–895. [PubMed] [Google Scholar]
  14. Liang L., Diehl-Jones W., Lasko P. Localization of vasa protein to the Drosophila pole plasm is independent of its RNA-binding and helicase activities. Development. 1994 May;120(5):1201–1211. doi: 10.1242/dev.120.5.1201. [DOI] [PubMed] [Google Scholar]
  15. Linder P., Lasko P. F., Ashburner M., Leroy P., Nielsen P. J., Nishi K., Schnier J., Slonimski P. P. Birth of the D-E-A-D box. Nature. 1989 Jan 12;337(6203):121–122. doi: 10.1038/337121a0. [DOI] [PubMed] [Google Scholar]
  16. Mattaj I. W. RNA recognition: a family matter? Cell. 1993 Jun 4;73(5):837–840. doi: 10.1016/0092-8674(93)90265-r. [DOI] [PubMed] [Google Scholar]
  17. Moazed D., Noller H. F. Interaction of tRNA with 23S rRNA in the ribosomal A, P, and E sites. Cell. 1989 May 19;57(4):585–597. doi: 10.1016/0092-8674(89)90128-1. [DOI] [PubMed] [Google Scholar]
  18. Moazed D., Noller H. F. Intermediate states in the movement of transfer RNA in the ribosome. Nature. 1989 Nov 9;342(6246):142–148. doi: 10.1038/342142a0. [DOI] [PubMed] [Google Scholar]
  19. Moazed D., Noller H. F. Sites of interaction of the CCA end of peptidyl-tRNA with 23S rRNA. Proc Natl Acad Sci U S A. 1991 May 1;88(9):3725–3728. doi: 10.1073/pnas.88.9.3725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Nishi K., Morel-Deville F., Hershey J. W., Leighton T., Schnier J. An eIF-4A-like protein is a suppressor of an Escherichia coli mutant defective in 50S ribosomal subunit assembly. Nature. 1988 Dec 1;336(6198):496–498. doi: 10.1038/336496a0. [DOI] [PubMed] [Google Scholar]
  21. Noller H. F. Ribosomal RNA and translation. Annu Rev Biochem. 1991;60:191–227. doi: 10.1146/annurev.bi.60.070191.001203. [DOI] [PubMed] [Google Scholar]
  22. Noller H. F. Structure of ribosomal RNA. Annu Rev Biochem. 1984;53:119–162. doi: 10.1146/annurev.bi.53.070184.001003. [DOI] [PubMed] [Google Scholar]
  23. Predki P. F., Nayak L. M., Gottlieb M. B., Regan L. Dissecting RNA-protein interactions: RNA-RNA recognition by Rop. Cell. 1995 Jan 13;80(1):41–50. doi: 10.1016/0092-8674(95)90449-2. [DOI] [PubMed] [Google Scholar]
  24. Rhoads R. E. Cap recognition and the entry of mRNA into the protein synthesis initiation cycle. Trends Biochem Sci. 1988 Feb;13(2):52–56. doi: 10.1016/0968-0004(88)90028-x. [DOI] [PubMed] [Google Scholar]
  25. Rozen F., Edery I., Meerovitch K., Dever T. E., Merrick W. C., Sonenberg N. Bidirectional RNA helicase activity of eucaryotic translation initiation factors 4A and 4F. Mol Cell Biol. 1990 Mar;10(3):1134–1144. doi: 10.1128/mcb.10.3.1134. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Schmid S. R., Linder P. D-E-A-D protein family of putative RNA helicases. Mol Microbiol. 1992 Feb;6(3):283–291. doi: 10.1111/j.1365-2958.1992.tb01470.x. [DOI] [PubMed] [Google Scholar]
  27. Tabor S., Richardson C. C. A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1074–1078. doi: 10.1073/pnas.82.4.1074. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Toone W. M., Rudd K. E., Friesen J. D. deaD, a new Escherichia coli gene encoding a presumed ATP-dependent RNA helicase, can suppress a mutation in rpsB, the gene encoding ribosomal protein S2. J Bacteriol. 1991 Jun;173(11):3291–3302. doi: 10.1128/jb.173.11.3291-3302.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES