C-13 NMR chemical shifts can predict disulfide bond formation

The presence of disulfide bonds can be detected unambiguously only by X-ray crystallography, and otherwise must be inferred by chemical methods. In th is study we demonstrate that C-13 NMR chemical shifts are diagnostic of dis ulfide bond formation, and can discriminate between cysteine in the reduced (free) and oxidized (disulfide bonded) state. A database of cysteine C-13 C-alpha and C-beta chemical shifts was constructed from the BMRB and Sheffi eld databases, and published journals. Statistical analysis indicated that the C-beta shift is extremely sensitive to the redox state, and can predict the disulfide-bonded state. Further, chemical shifts in both states occupy distinct clusters as a function of secondary structure in the C-alpha/C-be ta chemical shift map. On the basis of these results, we provide simple gro und rules for predicting the redox state of cysteines; these rules could be used effectively in NMR structure determination, predicting new folds, and in protein folding studies.