De novo protein design and electrochemistry were used to measure changes in the potential and pKA of a phenol when its OH group is moved from a solvent-exposed to a sequestered protein position. A "phenol rotation strategy" was adopted in which phenols, containing a SH in position 4, 3, or 2 relative to the OH group, were bound to a buried protein site. The α3C protein used here is a tryptophan to cysteine variant of the structurally defined α3W protein (Dai et al. (2002) J. Am. Chem. Soc. 124, 10952-10953). The protein characteristics of α3C and the three mercaptophenol-α3C (MP-α3C) proteins are shown to be close to those of α3W. Moreover, the phenol OH group is fully solvent exposed in 4MP-α3C and more sequestered in 3MP-α3C and 2MP-α3C. Here we compare the redox properties of the three mercaptophenols when bound to α3C and to cysteine free in water. The pKA and Epeak values are essential identical when 4MP is ligated to α3C relative to when it is free in solution. In contrast, these values are increased in 3MP-α3C and 2MP-α3C relative to the solvated compounds. The E peak vs pH plots all display a ~∼9 mV/pH unit dependence. We conclude that interactions with the OH group dominate the phenol redox characteristics. In 3MP-α3C and 2MP-α3C, hydrogen bonds between the protein and the bound phenols appear to either stabilize the reduced phenol or destabilize the radical, relative to the aqueous buffer, raising the potential by 0.11 and 0.12 V, respectively. © 2005 American Chemical Society.