The four lowest-energy isomers are depicted in Figure . Structure D's spectrum reproduces the new peak above 3500 cm -1 . Only structure D features a unique bonding arrangement in which one bisulfate S=O group accepts two hydrogen bonds: one from the second bisulfate O-H, and another from the carboxylic O-H of a G. The hydrogen bond between the two bisulfate groups is the shortest hydrogen bond in the structure, with a bond length of 1.66 Å. The sharp, intense transition at ∼ 3600 cm -1 in the calculated spectrum corresponds to the coupled stretching motion of this unique three way interaction. However, the complex pattern in this region also raises the possibility of contributions from more than one isomer. We obtained the spectrum of the second isomer (shown in Figure ) using hole burning to remove the population of the isomer that absorbs at 3505 cm -1 . The peaks between 3400 -3452 cm -1 in the experimental spectrum originate from the two different isomers, respectively. Boltzmann population analysis, as described in the SI, was conducted at both D 2 and N 2 tagging temperatures (47 K and 17 K, respectively, as shown in Figure ). Neglecting the contribution of the tag, the analysis revealed that the two isomers are close in energy, with a difference of approximately 0.03 -0.06 kcal mol -1 .