CYSTEINE CONFORMATION AND SULFHYDRYL INTERACTIONS IN PROTEINS AND VIRUSES .3. QUANTITATIVE MEASUREMENT OF THE RAMAN S-H BAND INTENSITY AND FREQUENCY

The bond stretching vibration of the cysteine sulfhydryl (SH) group in a typical protein generates a Raman band in the spectral interval 250 0-2600 cm-1, a region devoid of interference from any other fundamenta l mode of vibration of the protein. The relatively high Raman cross se ction associated with the S-H stretching vibration, the sensitivity of the vibrational frequency to hydrogen bonding interactions and side c hain configurations, and the dependence of the Raman intensity on thio l-thiolate equilibria, combine to make the Raman SH band a potentially valuable marker of protein sulfhydryl interactions and a unique indic ator of sulfhydryl participation in thiol-disulfide oxidoreductase act ivity. In order to exploit Raman spectroscopy for these purposes, accu rate and precise measurements of Raman SH band profiles are required. We show here that the required precision and accuracy can be achieved by use of the Raman band corresponding to the stretching vibration of in situ nitrogen gas as a quantitative intensity and frequency standar d. The Raman 0-branch center of the N2 band occurs at 2330.7 cm-1.