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 Mg(2+) and these nitrogen atoms are inherently difficult to observe in large RNAs. Rather than using the insensitive direct (15)N detection, here we have used (2)J-[(1)H,(15)N]-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.ai5gamma as an example, we show that direct N7 coordination of a Mg(2+) ion takes place in a tetraloop nucleotide. A second Mg(2+) 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.

• PMID: 20170966
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Erat MC, Kovacs H, Sigel RK

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