Fluorescence Characterization of the Transfer RNA-like Domain of Transfer Messenger RNA in Complex with Small Binding Protein B

Overview

Journal Biochemistry

Specialty Biochemistry

Date 2012 Apr 10

PMID 22482838

Citations 5

Authors

May Daher

David Rueda

Affiliations

Soon will be listed here.

Abstract

Transfer messenger RNA (tmRNA) and small binding protein B (SmpB) are the main components of the trans-translation rescue machinery that releases stalled ribosomes from defective mRNAs. Little is known about how SmpB binding affects the conformation of the tRNA-like domain (TLD) of tmRNA. It has been previously hypothesized that the absence of a D stem in the TLD provides flexibility in the elbow region of tmRNA, which can be stabilized by its interaction with SmpB. Here, we have used Förster resonance energy transfer to characterize the global structure of the tRNA-like domain of tmRNA in the presence and absence of SmpB and as a function of Mg(2+) concentration. Our results show tight and specific binding of SmpB to tmRNA. Surprisingly, our data show that the global conformation and flexibility of tmRNA do not change upon SmpB binding. However, Mg(2+) ions induce an 11 Å compaction in the tmRNA structure, suggesting that the flexibility in the H2a stem may allow different conformations of tmRNA as the TLD and mRNA-like domain need to be positioned differently while moving through the ribosome.

Citing Articles

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References

Miller M, Liu Z, Cazier D, Gebhard G, Herron S, Zaher H . The role of SmpB and the ribosomal decoding center in licensing tmRNA entry into stalled ribosomes. RNA. 2011; 17(9):1727-36. PMC: 3162337. DOI: 10.1261/rna.2821711. View

Keiler K . Biology of trans-translation. Annu Rev Microbiol. 2008; 62:133-51. DOI: 10.1146/annurev.micro.62.081307.162948. View

Weis F, Bron P, Rolland J, Thomas D, Felden B, Gillet R . Accommodation of tmRNA-SmpB into stalled ribosomes: a cryo-EM study. RNA. 2009; 16(2):299-306. PMC: 2811659. DOI: 10.1261/rna.1757410. View

Rueda D, Walter N . Fluorescent energy transfer readout of an aptazyme-based biosensor. Methods Mol Biol. 2006; 335:289-310. DOI: 10.1385/1-59745-069-3:289. View

Ivanova N, Pavlov M, Felden B, Ehrenberg M . Ribosome rescue by tmRNA requires truncated mRNAs. J Mol Biol. 2004; 338(1):33-41. DOI: 10.1016/j.jmb.2004.02.043. View

Rueda D, Hsieh J, Day-Storms J, Fierke C, Walter N . The 5' leader of precursor tRNAAsp bound to the Bacillus subtilis RNase P holoenzyme has an extended conformation. Biochemistry. 2005; 44(49):16130-9. DOI: 10.1021/bi0519093. View

Walter F, Putz J, Giege R, Westhof E . Binding of tobramycin leads to conformational changes in yeast tRNA(Asp) and inhibition of aminoacylation. EMBO J. 2002; 21(4):760-8. PMC: 125865. DOI: 10.1093/emboj/21.4.760. View

Zwieb C, Wower I, Wower J . Comparative sequence analysis of tmRNA. Nucleic Acids Res. 1999; 27(10):2063-71. PMC: 148424. DOI: 10.1093/nar/27.10.2063. View

Walter N . Probing RNA structural dynamics and function by fluorescence resonance energy transfer (FRET). Curr Protoc Nucleic Acid Chem. 2008; Chapter 11:11.10.1-11.10.23. DOI: 10.1002/0471142700.nc1110s11. View

10.

Jones C, Spencer A, Hsu J, Spremulli L, Martinis S, Derider M . A counterintuitive Mg2+-dependent and modification-assisted functional folding of mitochondrial tRNAs. J Mol Biol. 2006; 362(4):771-86. PMC: 1781928. DOI: 10.1016/j.jmb.2006.07.036. View

11.

Gutmann S, Haebel P, Metzinger L, Sutter M, Felden B, Ban N . Crystal structure of the transfer-RNA domain of transfer-messenger RNA in complex with SmpB. Nature. 2003; 424(6949):699-703. DOI: 10.1038/nature01831. View

12.

Hsieh J, Koutmou K, Rueda D, Koutmos M, Walter N, Fierke C . A divalent cation stabilizes the active conformation of the B. subtilis RNase P x pre-tRNA complex: a role for an inner-sphere metal ion in RNase P. J Mol Biol. 2010; 400(1):38-51. PMC: 2939038. DOI: 10.1016/j.jmb.2010.04.050. View

13.

Eis P, Millar D . Conformational distributions of a four-way DNA junction revealed by time-resolved fluorescence resonance energy transfer. Biochemistry. 1993; 32(50):13852-60. DOI: 10.1021/bi00213a014. View

14.

Sevenich F, Langowski J, Weiss V, Rippe K . DNA binding and oligomerization of NtrC studied by fluorescence anisotropy and fluorescence correlation spectroscopy. Nucleic Acids Res. 1998; 26(6):1373-81. PMC: 147426. DOI: 10.1093/nar/26.6.1373. View

15.

Pereira M, Harris D, Rueda D, Walter N . Reaction pathway of the trans-acting hepatitis delta virus ribozyme: a conformational change accompanies catalysis. Biochemistry. 2002; 41(3):730-40. DOI: 10.1021/bi011963t. View

16.

Dulebohn D, Choy J, Sundermeier T, Okan N, Karzai A . Trans-translation: the tmRNA-mediated surveillance mechanism for ribosome rescue, directed protein degradation, and nonstop mRNA decay. Biochemistry. 2007; 46(16):4681-93. DOI: 10.1021/bi6026055. View

17.

Himeno H, Sato M, Tadaki T, Fukushima M, Ushida C, Muto A . In vitro trans translation mediated by alanine-charged 10Sa RNA. J Mol Biol. 1997; 268(5):803-8. DOI: 10.1006/jmbi.1997.1011. View

18.

Lamichhane R, Daubner G, Thomas-Crusells J, Auweter S, Manatschal C, Austin K . RNA looping by PTB: Evidence using FRET and NMR spectroscopy for a role in splicing repression. Proc Natl Acad Sci U S A. 2010; 107(9):4105-10. PMC: 2840148. DOI: 10.1073/pnas.0907072107. View

19.

Klostermeier D, Millar D . Time-resolved fluorescence resonance energy transfer: a versatile tool for the analysis of nucleic acids. Biopolymers. 2002; 61(3):159-79. DOI: 10.1002/bip.10146. View

20.

Rueda D, Wick K, McDowell S, Walter N . Diffusely bound Mg2+ ions slightly reorient stems I and II of the hammerhead ribozyme to increase the probability of formation of the catalytic core. Biochemistry. 2003; 42(33):9924-36. DOI: 10.1021/bi0347757. View