Revealing weak histidine ¹⁵N homonuclear scalar couplings using Solid-State Magic-Angle-Spinning NMR spectroscopy.

ABSTRACT: The tautomeric structure and chemistry of the histidine imidazole ring play active roles in many structurally and functionally important proteins and polypeptides. While in NMR spectroscopy histidine chemical shifts (e.g. ¹⁵N, ¹³C, and ¹H) have been commonly used to characterize the tautomeric structure, hydrogen bonding, and torsion angles, homonuclear ¹⁵N scalar couplings in histidine have rarely been reported. Here, we propose double spin-echo sequences to compare the observed signals with and without a 90° pulse between the two spin-echo periods, such that their signal ratio as a function of the echo time solely depends on homonuclear scalar couplings, allowing for measuring weak homonuclear scalar couplings without influence from transverse dephasing effects, thus capable of revealing hydrogen-bond mediated ¹⁵N-¹⁵N J-couplings that can provide direct and definitive evidence for the formation of N^…H^…N hydrogen-bonding associated with the imidazole ring. We used two ¹³C,¹⁵N labeled histidine samples recrystallized from solutions at pH 6.3 and pH 11.0 to demonstrate the feasibility of this method and reveal the existence of a weak two-bond scalar coupling between the N_δ1 and N_ε2 sites in the histidine imidazole ring in three tautomeric states and the presence of a hydrogen-bond mediated scalar coupling between the N_δ1 site in the imidazole ring and the backbone N_α site in the histidine neutral τ and π states. Our results demonstrate that weak ¹⁵N homonuclear scalar couplings can be measured even when their values are less than their corresponding intrinsic natural linewidths, thus providing direct and definitive evidence for the formation of N^…H^…N hydrogen bonding that is associated with the histidine imidazole ring.

SUBMITTER: Tan C 

PROVIDER: S-EPMC7426724 | biostudies-literature |

REPOSITORIES: biostudies-literature