Hydrostatic pressure response of α-keratin investigated with Raman spectroscopy

Keratins represent an important class of sulfur-rich structural proteins. In this study, the pressure response of α-keratin, extracted from sheep wool, was investigated using Raman spectroscopy up to 4 GPa. A non-polar liquid (a Fluorinert™ FC70/77 mixture) served as the Pressure Transmitting Medium (PTM) in a Diamond Anvil Cell (DAC). Scanning Electron Microscopy (SEM) and Small-Angle X-ray Scattering (SAXS) were used for the morphological and structural characterization of the sample. Three Raman bands were monitored during compression and decompression: CH deformation, amide-I (dominated by the CO stretching vibration of the peptide bond) and CH stretching vibrations centered at approximately 1450, 1670 and 2940 cm^-1, respectively. The deformation modes of the CH bonds exhibited a weak positive pressure dependence (∼1.7 cm^-1/GPa), whereas the corresponding stretching modes showed a much stronger dependence (6-9 cm^-1/GPa). A positive slope (∼2.2 cm^-1/GPa) was also observed for the amide-I band, consistent with CO strengthening in the peptide bonds under compression. This behavior contrasts with that of collagen, which exhibits a negative pressure slope for the same vibrational mode. The opposite responses are discussed in relation to the different structural roles of hydrogen bonding in these two fibrous proteins. Overall, the response of keratin to high hydrostatic pressure is reversible.