A 2D Nuclear Overhauser Effect Spectroscopy (NOESY) spectrum highlights neighboring base pairs observed by NMR, with blue lines referencing off-diagonal peaks, along with corresponding assignments (Figure ). Interestingly, off-diagonal peaks for nucleotides neighboring the 2×2 internal loop, G4 and G18, are broadened due to exchange between multiple structures, and do not show any NOEs to the internal loop resonances (Figure ). In addition, there are two sets of off-diagonal peaks in the non-canonical region that do not have NOEs to either the upper or lower helices, which are highlighted with red boxes and appear between 10 and 12 ppm (Figure ). We suspected that these peaks corresponded to specific structure(s) in the 2×2 GG/GG internal loop but could not make definitive assignments due to exchange broadening of these dynamic nucleotides observed in the NMR spectra (Figure ).