Abstract
The four aromatic amino acids in proteins, namely histidine, phenylalanine, tyrosine, and tryptophan, have strongly overlapping 13C chemical shift ranges between 100 and 160ppm, and have so far been largely neglected in solid-state NMR determination of protein structures. Yet aromatic residues play important roles in biology through π–π and cation–π interactions. To better resolve and assign aromatic residues' 13C signals in magic-angle-spinning (MAS) solid-state NMR spectra, we introduce two spectral editing techniques. The first method uses gated 1H decoupling in a proton-driven spin-diffusion (PDSD) experiment to remove all protonated 13C signals and retain only non-protonated carbon signals in the aromatic region of the 13C spectra. The second technique uses chemical shift filters and 1H–13C dipolar dephasing to selectively detect the Cα, Cβ and CO cross peaks of aromatic residues while suppressing the signals of all aliphatic residues. We demonstrate these two techniques on amino acids, a model peptide, and the microcrystalline protein GB1, and show that they significantly simplify the 2D NMR spectra and both reveal and permit the ready assignment of the aromatic residues' signals.
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•Two methods are described to selectively detect the signals of aromatic residues.•Gated 1H decoupling selectivelydetects unprotonated aromatic 13C signals.•Chemical shift filters select the Cα, Cβ and CO signals of aromatic residues.•We successfully demonstrated these techniques on model peptides and proteins.