A (13)C-detected (15)N double-quantum NMR experiment to probe arginine side-chain guanidinium (15)N(η) chemical shifts

Arginine side-chains are often key for enzyme catalysis, protein–ligand and protein–protein interactions. The importance of arginine stems from the ability of the terminal guanidinium group to form many key interactions, such as hydrogen bonds and salt bridges, as well as its perpetual positive charge. We present here an arginine (13)C(ζ)-detected NMR experiment in which a double-quantum coherence involving the two (15)N(η) nuclei is evolved during the indirect chemical shift evolution period. As the precession frequency of the double-quantum coherence is insensitive to exchange of the two (15)N(η); this new approach is shown to eliminate the previously deleterious line broadenings of (15)N(η) resonances caused by the partially restricted rotation about the C(ζ)–N(ε) bond. Consequently, sharp and well-resolved (15)N(η) resonances can be observed. The utility of the presented method is demonstrated on the L99A mutant of the 19 kDa protein T4 lysozyme, where the measurement of small chemical shift perturbations, such as one-bond deuterium isotope shifts, of the arginine amine (15)N(η) nuclei becomes possible using the double-quantum experiment. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s10858-017-0137-2) contains supplementary material, which is available to authorized users.