Terahertz dielectric response sensitivity to protein oxidation state

Previously we have shown that the terahertz dielectric response is sensitive to the oxidation state for cytochrome c (CytC) films. Here we discuss the origin of this sensitivity through hydration dependent measurements on films, solution phase measurements as a function of temperature and normal mode calculations as a function of hydration. We find that the hydration dependence of the terahertz response rapidly approaches its fully solvated value for ferri CytC, whereas for ferro CytC the effect of added water is more gradual, with the system not reaching fully hydrated values up to 0.5 gm water/gm protein. For solution phase samples below 270 K we do not see a significant difference in the terahertz response suggesting that the fully hydrated ferro cytochrome c has the same picosecond dynamics as ferri cytochrome c. These results contradict X-ray B factor measurements that suggest that ferri cytochrome c is significantly more flexible than ferro suggesting that the B-factor determination from X-ray crystal measurements may not represent in vivo values as crystal water is less than 0.5 gm water/gm protein.