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. 2002 Feb;8(2):252–260. doi: 10.1017/s1355838202014826

Water counting: quantitating the hydration level of paramagnetic metal ions bound to nucleotides and nucleic acids.

Charles G Hoogstraten 1, R David Britt 1
PMCID: PMC1370247  PMID: 11911370

Abstract

Binding of divalent metal ions plays a key role in the structure and function of ribozymes and other RNAs. In turn, the energetics and kinetics of the specific binding process are dominated by the balance between the cost of dehydrating the aqueous ion and the energy gained from inner-sphere interactions with the macromolecule. In this work, we introduce the use of the pulsed EPR technique of 2H Electron Spin-Echo Envelope Modulation (ESEEM) to determine the hydration level of Mn2+ ions bound to nucleotides and nucleic acids. Mn2+ is an excellent structural and functional mimic for Mg2+, the most common divalent ion of physiological interest. Comparison of data in D2O and H2O, with aqueous Mn2+ as a reference standard, allows a robust and precise determination of the number of bound water molecules, and therefore the number of RNA-derived ligands. Examples of applications to the mononucleotide models MnGMP and MnATP, as well as to the paradigmatic RNA system tRNAPhe, are shown.

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Selected References

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  1. Bellew B. F., Halkides C. J., Gerfen G. J., Griffin R. G., Singel D. J. High frequency (139.5 GHz) electron paramagnetic resonance characterization of Mn(II)-H2(17)O interactions in GDP and GTP forms of p21 ras. Biochemistry. 1996 Sep 17;35(37):12186–12193. doi: 10.1021/bi960594b. [DOI] [PubMed] [Google Scholar]
  2. Bernat B. A., Laughlin L. T., Armstrong R. N. Fosfomycin resistance protein (FosA) is a manganese metalloglutathione transferase related to glyoxalase I and the extradiol dioxygenases. Biochemistry. 1997 Mar 18;36(11):3050–3055. doi: 10.1021/bi963172a. [DOI] [PubMed] [Google Scholar]
  3. De Meester P., Goodgame D. M., Jones T. J., Skapski A. C. X-ray evidence for metal-N-7 bonding in a hydrated manganese derivative of guanosine 5'-monophosphate. Biochem J. 1974 Jun;139(3):791–792. doi: 10.1042/bj1390791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Draper D. E., Misra V. K. RNA shows its metal. Nat Struct Biol. 1998 Nov;5(11):927–930. doi: 10.1038/2901. [DOI] [PubMed] [Google Scholar]
  5. Draper D. E. On the coordination properties of Eu3+ bound to tRNA. Biophys Chem. 1985 Feb;21(2):91–101. doi: 10.1016/0301-4622(85)85011-0. [DOI] [PubMed] [Google Scholar]
  6. Feig A. L., Panek M., Horrocks W. D., Jr, Uhlenbeck O. C. Probing the binding of Tb(III) and Eu(III) to the hammerhead ribozyme using luminescence spectroscopy. Chem Biol. 1999 Nov;6(11):801–810. doi: 10.1016/s1074-5521(99)80127-6. [DOI] [PubMed] [Google Scholar]
  7. Feig A. L., Scott W. G., Uhlenbeck O. C. Inhibition of the hammerhead ribozyme cleavage reaction by site-specific binding of Tb. Science. 1998 Jan 2;279(5347):81–84. doi: 10.1126/science.279.5347.81. [DOI] [PubMed] [Google Scholar]
  8. Feigon J., Butcher S. E., Finger L. D., Hud N. V. Solution nuclear magnetic resonance probing of cation binding sites on nucleic acids. Methods Enzymol. 2001;338:400–420. doi: 10.1016/s0076-6879(02)38230-2. [DOI] [PubMed] [Google Scholar]
  9. Gonzalez R. L., Jr, Tinoco I., Jr Identification and characterization of metal ion binding sites in RNA. Methods Enzymol. 2001;338:421–443. doi: 10.1016/s0076-6879(02)38231-4. [DOI] [PubMed] [Google Scholar]
  10. Halkides C. J., Farrar C. T., Singel D. J. The effects of cryoprotection on the structure and activity of p21 ras: implications for electron spin-echo envelope modulation spectroscopy. J Magn Reson. 1998 Sep;134(1):142–153. doi: 10.1006/jmre.1998.1520. [DOI] [PubMed] [Google Scholar]
  11. Hurd R. E., Azhderian E., Reid B. R. Paramagnetic ion effects on the nuclear magnetic resonance spectrum of transfer ribonucleic acid: assignment of the 15--48 tertiary resonance. Biochemistry. 1979 Sep 4;18(18):4012–4017. doi: 10.1021/bi00585a026. [DOI] [PubMed] [Google Scholar]
  12. Jack A., Ladner J. E., Rhodes D., Brown R. S., Klug A. A crystallographic study of metal-binding to yeast phenylalanine transfer RNA. J Mol Biol. 1977 Apr 15;111(3):315–328. doi: 10.1016/s0022-2836(77)80054-5. [DOI] [PubMed] [Google Scholar]
  13. Latwesen D. G., Poe M., Leigh J. S., Reed G. H. Electron paramagnetic resonance studies of a ras p21-MnIIGDP complex in solution. Biochemistry. 1992 Jun 2;31(21):4946–4950. doi: 10.1021/bi00136a004. [DOI] [PubMed] [Google Scholar]
  14. Leroy J. L., Guéron M. Electrostatic effects in divalent ion binding to tRNA. Biopolymers. 1977 Nov;16(11):2429–2446. doi: 10.1002/bip.1977.360161108. [DOI] [PubMed] [Google Scholar]
  15. Misra V. K., Draper D. E. A thermodynamic framework for Mg2+ binding to RNA. Proc Natl Acad Sci U S A. 2001 Oct 23;98(22):12456–12461. doi: 10.1073/pnas.221234598. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Misra V. K., Draper D. E. Mg(2+) binding to tRNA revisited: the nonlinear Poisson-Boltzmann model. J Mol Biol. 2000 Jun 9;299(3):813–825. doi: 10.1006/jmbi.2000.3769. [DOI] [PubMed] [Google Scholar]
  17. Misra V. K., Draper D. E. On the role of magnesium ions in RNA stability. Biopolymers. 1998;48(2-3):113–135. doi: 10.1002/(SICI)1097-0282(1998)48:2<113::AID-BIP3>3.0.CO;2-Y. [DOI] [PubMed] [Google Scholar]
  18. Prisner T., Rohrer M., MacMillan F. Pulsed EPR spectroscopy: biological applications. Annu Rev Phys Chem. 2001;52:279–313. doi: 10.1146/annurev.physchem.52.1.279. [DOI] [PubMed] [Google Scholar]
  19. Pyle A. M. Ribozymes: a distinct class of metalloenzymes. Science. 1993 Aug 6;261(5122):709–714. doi: 10.1126/science.7688142. [DOI] [PubMed] [Google Scholar]
  20. Reed G. H., Poyner R. R. Mn2+ as a probe of divalent metal ion binding and function in enzymes and other proteins. Met Ions Biol Syst. 2000;37:183–207. [PubMed] [Google Scholar]
  21. Schimmel P. R., Redfield A. G. Transfer RNA in solution: selected topics. Annu Rev Biophys Bioeng. 1980;9:181–221. doi: 10.1146/annurev.bb.09.060180.001145. [DOI] [PubMed] [Google Scholar]
  22. Serpersu E. H., McCracken J., Peisach J., Mildvan A. S. Electron spin echo modulation and nuclear relaxation studies of staphylococcal nuclease and its metal-coordinating mutants. Biochemistry. 1988 Oct 18;27(21):8034–8044. doi: 10.1021/bi00421a010. [DOI] [PubMed] [Google Scholar]
  23. Shi H., Moore P. B. The crystal structure of yeast phenylalanine tRNA at 1.93 A resolution: a classic structure revisited. RNA. 2000 Aug;6(8):1091–1105. doi: 10.1017/s1355838200000364. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Sigel H. Isomeric equilibria in complexes of adenosine 5'-triphosphate with divalent metal ions. Solution structures of M(ATP)2- complexes. Eur J Biochem. 1987 May 15;165(1):65–72. doi: 10.1111/j.1432-1033.1987.tb11194.x. [DOI] [PubMed] [Google Scholar]
  25. Stein A., Crothers D. M. Equilibrium binding of magnesium(II) by Escherichia coli tRNAfMet. Biochemistry. 1976 Jan 13;15(1):157–160. doi: 10.1021/bi00646a024. [DOI] [PubMed] [Google Scholar]
  26. Walters J. A., Geerdes H. A., Hilbers C. W. On the binding of Mg2+ and Mn2+ to tRNA. Biophys Chem. 1977 Sep;7(2):147–151. doi: 10.1016/0301-4622(77)80007-0. [DOI] [PubMed] [Google Scholar]

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