2-DIMENSIONAL NMR EXPERIMENTS FOR THE ASSIGNMENT OF AROMATIC SIDE-CHAINS IN C-13-LABELED PROTEINS

As aromatic residues very often are part of the hydrophobic core of pr oteins, the unambiguous assignment of the aromatic proton resonances i s essential for an accurate and precise structure determination. Inste ad of transferring H-1(beta) coherence to the aromatic protons via C-1 3(gamma) like in a number of published methods, in our new experiments the C-13(gamma) resonances are first correlated with the H-1(beta) ch emical shifts in one experiment and then with the aromatic proton reso nances in four other experiments, Their short coherence transfer pathw ays make the experiments applicable to proteins with a molecular weigh t larger than 20 kDa, as is demonstrated for Fusarium solani pisi cuti nase (214 residues), The dispersion of the C-gamma chemical shifts bet ween different aromatic residue types is obvious, but even the dispers ion within one type is sufficient to combine the experiments using onl y the C-gamma chemical shift and to assign nearly all aromatic proton resonances of cutinase. (C) 1998 Academic Press.