Identification of the Metal Ion Binding Site on an RNA Motif from Hammerhead Ribozymes Using <sup>15</sup>N NMR Spectroscopy

Tanaka, Yoshiyuki; Kojima, Chojiro; Morita, Eugene Hayato; Kasai, Yusuke; Yamasaki, Kazuhiko; Ono, Akira; Kainosho, Masatsune; Taira, Kazunari

doi:10.1021/ja011520c

Cited by 49 publications

(54 citation statements)

References 23 publications

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“…[21] The proton exchange rate between the Ag + -free and Ag + -bound duplex DNAs was slow relative to the timescale of the NMR measurement, [21] although the exchange rates of metal ion association-dissociation processes with DNA/RNA molecules are usually fast relative to the timescale of the NMR measurement. [40] A similar slow proton exchange rate was also observed in the 1 H NMR study of T-Hg-T base pair formation, where the Hg 2+ ion bound to the N3 positions of two thymine bases. [19] Thus, the observed slow proton exchange rate for C-Ag-C formation suggests that the Ag + ion may bind to the inner N3 positions rather than to the outer O2 or N4 positions.…”

Section: Discussionsupporting

confidence: 61%

Thermodynamic Properties of the Specific Binding Between Ag⁺Ions and C:C Mismatched Base Pairs in Duplex DNA

Torigoe

Miyakawa

Ono

et al. 2011

Nucleosides, Nucleotides and Nucleic Acids

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Metal-mediated base pairs formed by the interaction between metal ions and artificial bases in oligonucleotides have been developed for potential applications in nanotechnology. We recently found that a natural C:C mismatched base pair bound to an Ag(+) ion to generate a novel metal-mediated base pair in duplex DNA. Preparation of the novel C-Ag-C base pair involving natural bases is more convenient than that of metal-mediated base pairs involving artificial bases because time-consuming base synthesis is not required. Here, we examined the thermodynamic properties of the binding between the Ag(+) ion and each of single and double C:C mismatched base pair in duplex DNA by isothermal titration calorimetry. The Ag(+) ion specifically bound to the C:C mismatched base pair at a 1:1 molar ratio with 10(6) M(-1) binding constant, which was significantly larger than those for nonspecific metal ion-DNA interactions. The specific binding between the Ag(+) ion and the single C:C mismatched base pair was mainly driven by the positive dehydration entropy change and the negative binding enthalpy change. In the interaction between the Ag(+) ion and each of the consecutive and interrupted double C:C mismatched base pairs, stoichiometric binding at a 1:1 molar ratio was achieved in each step of the first and second Ag(+) binding. The binding affinity for the second Ag(+) binding was similar to that for the first Ag(+) binding. Stoichiometric binding without interference and negative cooperativity may be favorable for aligning multiple Ag(+) ions in duplex DNA for applications of the metal-mediated base pairs in nanotechnology.

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Section: Discussionsupporting

confidence: 61%

Thermodynamic Properties of the Specific Binding Between Ag⁺Ions and C:C Mismatched Base Pairs in Duplex DNA

Torigoe

Miyakawa

Ono

et al. 2011

Nucleosides, Nucleotides and Nucleic Acids

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“…Also here, phosphorothioate studies helped to prove the coordination of divalent metal ions during catalysis [271,272]. resonance, respectively [274][275][276][277]. The catalytic activity of the hammerhead ribozyme was also shown to be in principle independent of divalent metal ions [278] 113 Cd NMR, which allow to observe both the RNA liganding sites and the metal ion simultaneously.…”

Section: Cadmium(ii)-rescue Experimentsmentioning

confidence: 86%

Complex Formation of Cadmium with Sugar Residues, Nucleobases, Phosphates, Nucleotides, and Nucleic Acids

Sigel

Skilandat

Sigel

et al. 2012

Metal Ions in Life Sciences

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Cadmium(II), commonly classified as a relatively soft metal ion, prefers indeed aromaticnitrogen sites (e.g., N7 of purines) over oxygen sites (like sugar-hydroxyl groups). However, matters are not that simple, though it is true that the affinity of Cd(2+) towards ribose-hydroxyl groups is very small; yet, a correct orientation brought about by a suitable primary binding site and a reduced solvent polarity, as it is expected to occur in a folded nucleic acid, may facilitate metal ion-hydroxyl group binding very effectively. Cd(2+) prefers the guanine(N7) over the adenine(N7), mainly because of the steric hindrance of the (C6)NH(2) group in the adenine residue. This Cd(2+)-(N7) interaction in a guanine moiety leads to a significant acidification of the (N1)H meaning that the deprotonation reaction occurs now in the physiological pH range. N3 of the cytosine residue, together with the neighboring (C2)O, is also a remarkable Cd(2+) binding site, though replacement of (C2)O by (C2)S enhances the affinity towards Cd(2+) dramatically, giving in addition rise to the deprotonation of the (C4)NH(2) group. The phosphodiester bridge is only a weak binding site but the affinity increases further from the mono-to the di-and the triphosphate. The same also holds for the corresponding nucleotides. Complex stability of the pyrimidine-nucleotides is solely determined by the coordination tendency of the phosphate group(s), whereas in the case of purine-nucleotides macrochelate formation takes place by the interaction of the phosphate-coordinated Cd(2+) with N7. The extents of the formation degrees of these chelates are summarized and the effect of a non-bridging sulfur atom in a thiophosphate group (versus a normal phosphate group) is considered. Mixed ligand complexes containing a nucleotide and a further mono-or bidentate ligand are covered and it is concluded that in these species N7 is released from the coordination sphere of Cd(2+). In the case that the other ligand contains an aromatic residue (e.g., 2,2'-bipyridine or the indole ring of tryptophanate) intramolecular stack formation takes place. With buffers like Tris or Bistris mixed ligand complexes are formed. Cd(2+) coordination to dinucleotides and to dinucleoside monophosphates provides some insights regarding the interaction between Cd(2+) and nucleic acids. Cd(2+) binding to oligonucleotides follows the principles of coordination to its units. The available crystal studies reveal that N7 of purines is the prominent binding site followed by phosphate oxygens and other heteroatoms in nucleic acids. Due to its high thiophilicity, Cd(2+) is regularly used in so-called thiorescue experiments, which lead to the identification of a direct involvement of divalent metal ions in ribozyme catalysis.

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“…5,6,[13][14][15] The ring nitrogens of the purine bases in the hammerhead ribozyme have been probed for M n+ binding by observing the change of 15 N chemical shift using direct 15 N detection. 16 Recently, the coordination of Hg 2+ to N3 of thymine has been proven by the same method by incorporating specifically labelled nucleotides into a DNA duplex. 17 However, due to its low gyromagnetic moment, direct 15 N detection is very time consuming and the assignment of the peaks in a 15 N spectrum of a large RNA rather complicated.…”

mentioning

confidence: 99%

Metal ion-N7 coordination in a ribozyme branch domain by NMR

Erat¹,

Kovacs²,

Sigel³

2010

Journal of Inorganic Biochemistry

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The N7 of purine nucleotides presents one of the most dominant metal ion binding sites in nucleic acids. However, the interactions between kinetically labile metal ions like Mg2+ and these nitrogen atoms are inherently difficult to observe in large RNAs. Rather than using the insensitive direct N-15 detection, here we have used (2)J-H-1,N-15]-HSQC (Heteronuclear Single Quantum Coherence) NMR experiments as a fast and efficient method to specifically observe and characterize such interactions within larger RNA constructs. Using the 27 nucleotides long branch domain of the yeast-mitochondrial group II intron ribozyme Sc.ai5 gamma as an example, we show that direct N7 coordination of a Mg2+ ion takes place in a tetraloop nucleotide. A second Mg2+ ion, located in the major groove at the catalytic branch site, coordinates mainly in an outer-sphere fashion to the highly conserved flanking GU wobble pairs but not to N7 of the sandwiched branch adenosine. (C)

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Identification of the Metal Ion Binding Site on an RNA Motif from Hammerhead Ribozymes Using ¹⁵N NMR Spectroscopy

Cited by 49 publications

References 23 publications

Thermodynamic Properties of the Specific Binding Between Ag⁺Ions and C:C Mismatched Base Pairs in Duplex DNA

Thermodynamic Properties of the Specific Binding Between Ag⁺Ions and C:C Mismatched Base Pairs in Duplex DNA

Complex Formation of Cadmium with Sugar Residues, Nucleobases, Phosphates, Nucleotides, and Nucleic Acids

Metal ion-N7 coordination in a ribozyme branch domain by NMR

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Identification of the Metal Ion Binding Site on an RNA Motif from Hammerhead Ribozymes Using 15N NMR Spectroscopy

Cited by 49 publications

References 23 publications

Thermodynamic Properties of the Specific Binding Between Ag+Ions and C:C Mismatched Base Pairs in Duplex DNA

Thermodynamic Properties of the Specific Binding Between Ag+Ions and C:C Mismatched Base Pairs in Duplex DNA

Complex Formation of Cadmium with Sugar Residues, Nucleobases, Phosphates, Nucleotides, and Nucleic Acids

Metal ion-N7 coordination in a ribozyme branch domain by NMR

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Identification of the Metal Ion Binding Site on an RNA Motif from Hammerhead Ribozymes Using ¹⁵N NMR Spectroscopy

Thermodynamic Properties of the Specific Binding Between Ag⁺Ions and C:C Mismatched Base Pairs in Duplex DNA

Thermodynamic Properties of the Specific Binding Between Ag⁺Ions and C:C Mismatched Base Pairs in Duplex DNA