Two-Dimensional NMR Experiments for the Assignment of Aromatic Side Chains in 13C-labeled Proteins.

As aromatic residues very often are part of the hydrophobic core of proteins, the unambiguous assignment of the aromatic proton resonances is essential for an accurate and precise structure determination. Instead of transferring 1Hbeta coherence to the aromatic protons via 13Cgamma like in a number of published methods, in our new experiments the 13Cgamma resonances are first correlated with the 1Hbeta chemical shifts in one experiment and then with the aromatic proton resonances in four other experiments. Their short coherence transfer pathways make the experiments applicable to proteins with a molecular weight larger than 20 kDa, as is demonstrated for Fusarium solani pisi cutinase (214 residues). The dispersion of the Cgamma chemical shifts between different aromatic residue types is obvious, but even the dispersion within one type is sufficient to combine the experiments using only the Cgamma chemical shift and to assign nearly all aromatic proton resonances of cutinase. Copyright 1998 Academic Press. Copyright 1998 Academic Press