The development of a probe for sample tubes of 20-mm outside diameter has increased the sensitivity of natural abundance 13C Fourier transform nuclear magnetic resonance to the point that single-carbon resonances of proteins can be studied. Numerous narrow single-carbon resonances are observed in the aromatic region of the 13C spectrum of native hen egg-white lysozyme. Theoretical and experimental evidence is presented to show that these narrow resonances are those of the 28 nonprotonated aromatic carbons. The 59 protonated aromatic carbons give rise to a background of broad peaks. Partial assignments for the nonprotonated carbon resonances are presented. Significant chemical-shift variations occur upon folding of the protein into its native conformation. For example, the γ carbons of the six tryptophan residues resonate at 81.4, 82.1, 83.2, 83.8, and 85.2 ppm upfield from CS2. The peak at 85.2 ppm is a two-carbon resonance. Upon denaturation with guanidinium chloride, all six carbons resonate at about 83.8 ppm. 13C chemical shifts and assignments for aqueous tryptophan at pH 4 are also presented.
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