Complex formation of the tetracycline‐binding aptamer investigated by specific cross‐relaxation under DNP

While dynamic nuclear polarization (DNP) under magic‐angle spinning (MAS) is generally a powerful method capable of greatly enhancing the sensitivity of solid‐state NMR, the displacement of nuclear spin polarization far from thermal equilibrium (i.e., hyperpolarization) also gives rise to peculiar spin dynamics. Here, we elucidate how Specific Cross‐Relaxation Enhancement by Active Motions under DNP (SCREAM‐DNP) can be utilized in order to selectively obtain MAS NMR spectra of an RNA aptamer in a tightly bound complex with a methyl‐bearing ligand (tetracycline) due to the effective CH₃‐reorientation at an optimized sample temperature of ~160 K. We demonstrate that SCREAM‐DNP is able to spectrally isolate the complex from non‐bound species within an RNA mixture and perform a competition assay between the aptamer and a mutant with compromised binding affinity. Finally, we discuss variations in molecular structure and methyl dynamics as observed by SCREAM‐DNP between free tetracycline and when bound in a complex with RNA.

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