Differentiation of histidine tautomeric states using 15N selectively filtered 13C solid-state NMR spectroscopy

The histidine imidazole ring in proteins usually contains a mixture of three possible tautomeric states (two neutral - tau and pi states and a charged state) at physiological pHs. Differentiating the tautomeric states is critical for understanding how the histidine residue participates in many structurally and functionally important proteins. In this work, one dimensional N-15 selectively filtered C-13 solid-state NMR spectroscopy is proposed to differentiate histidine tautomeric states and to identify all C-13 resonances of the individual imidazole rings in a mixture of tautomeric states. When N-15 selective 180 degrees pulses are applied to the protonated or non-protonated nitrogen region, the C-13 sites that are bonded to the non-protonated or protonated nitrogen sites can be identified, respectively. A sample of C-13, N-15 labeled histidine powder lyophilized from a solution at pH 6.3 has been used to illustrate the usefulness of this scheme by uniquely assigning resonances of the neutral tau and charged states from the mixture. (C) 2014 Elsevier Inc. All rights reserved.