Multiple-Quantum Coherence Dramatically Enhances the Sensitivity of CH and CH₂ Correlations in Uniformly ¹³C-Labeled RNA

Abstract

One-bond 1H,13C and geminal 1H,1H dipolar interactions are normally the dominant causes of 1H and 13C transverse relaxation in NMR experiments applied to 13C,15N-labeled RNA in solution. Proton,carbon multiple-quantum coherences, where all heteronuclei connected by single bonds are evolved simultaneously in the transverse plane, however, are not affected by these strong dipolar interactions. Consequently, the transverse lifetimes, or T2 values, of these resonances can be dramatically extended. Here, we show that pulse sequences can be written that take advantage of this effect to enhance the sensitivity with which CH and CH2 correlations are observed in uniformly 13C,15N-labeled RNA oligonucleotides. In heteronuclear multiple-quantum correlation experiments, CH and CH2 correlations are detected with a sensitivity that is enhanced by about a factor of 3 relative to heteronuclear, single-quantum experiments for a 13C,15N-labeled 36mer RNA oligonucleotide and a constant time period of 25 ms. By including 1H,13C multiple-quantum coherence steps in an 1H,13C,15N NMR experiment of the “out and back” type currently used for through-bond resonance assignment in RNA, we have obtained a sensitivity enhancement of about a factor of 5 in the same 36mer RNA.