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Sensitivity optimized HCN and HCNCH experiments for 13C/15N labeled oligonucleotides
Authors:Radovan Fiala  Feng Jiang  Vladimír Sklenář
Affiliation:(1) Laboratory of Biomolecular Structure & Dynamics, Masaryk University, Kotlá"rcaron"ská 2, CZ-611 37 Brno, Czech Republic;(2) Cellular Biochemistry and Biophysics Program, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY, 10021, U.S.A
Abstract:Triple resonance HCN and HCNCH experiments used in studies of 13C/15N labeled oligonucleotides include extended evolution periods (typically up to 100 ms) to allow coherence transfer through a complex heteronuclear spin network. Unfortunately, most of the magnetization is lost during the evolution due to fast spin–spin relaxation dominated by one-bond 1H–13C dipolar interaction. As demonstrated recently, the sensitivity of the experiments can be dramatically improved by keeping the spin system in a state of proton–carbon multiple-quantum coherence, which is not affected by the strong dipolar coupling. However, the multiple-quantum coherence is very sensitive to homonuclear as well as long-range heteronuclear interactions. Unwanted magnetization transfer due to these interactions can reduce the sensitivity back to the level of a single-quantum experiment and, for some spin moieties, even eliminate the signal completely. In the present paper we show that a modified HCN scheme that refocuses the interfering coherences improves sensitivity routinely by a factor of 1.5 to 4 over a nonselective experiment. In addition, novel multiple-quantum 2D and 3D HCNCH experiments with substantially enhanced sensitivity are presented.
Keywords:assignment  labeled RNA  oligonucleotide  triple resonance
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