CYSTEINE CONFORMATION AND SULFHYDRYL INTERACTIONS IN PROTEINS AND VIRUSES .1. CORRELATION OF THE RAMAN S-H BAND WITH HYDROGEN-BONDING AND INTRAMOLECULAR GEOMETRY IN MODEL COMPOUNDS

The cysteine sulfhydryl group plays an important role in structural biochemistry. Cysteine thiols of proteins are capable of donating and accepting hydrogen bonds with solvent molecules as well as with other protein groups, and cysteine ligand coordination is fundamental to enzyme activity and nucleic acid recognition. We have undertaken a systematic Raman study of model mercaptans and cysteine thiols to provide an effective spectroscopic probe of the S-H group and its biologically relevant configurations and interactions in aqueous and crystalline proteins. The present study of aliphatic and aromatic mercaptans in both polar and apolar solvents, and of L-cysteine and glutathione in the crystal, provides a basis for interpreting the S-H stretching region of the Raman spectrum in terms of hydrogen bond donation by S-H, hydrogen bond acceptance by S, and rotamer populations of the cysteinyl side chain. The most definitive change observed in the Raman S-H stretching band is its shift to lower frequency when S-H acts as a hydrogen bond donor. We find further that the frequency interval in which the Raman S-H stretching vibration (sigma-SH) occurs is diagnostic of S-H donors which are strongly hydrogen bonded (2525-2560 cm-1), moderately hydrogen bonded (2560-2575 cm-1), or weakly hydrogen bonded (2575-2580 cm-1). When the S-H group is essentially non-hydrogen-bonded e.g., at high dilution in CCl4, we find-sigma-SH almost-equal-to 2585 +/- 5 cm-1. On the other hand, hydrogen bond acceptance by S in the absence of S-H donation elevates sigma-SH slightly (< 4 cm-1). In model mercaptans, P(C) and P(H) rotamers with respect to the C-C-S-H torsion (chi-2) yield sigma-SH values that are separated by approximately 10 cm-1, with the P(H) rotamer generally exhibiting the higher frequency of the two. Since the S-H region of the Raman spectrum contains no interference from other molecular vibrations, the approximate 10-cm-1 shift should be measurable in proteins and thus should provide a means of resolving different cysteine side-chain orientations.