Thermodynamic basis for redox regulation of the Yap1 signal transduction pathway

摘要

The Yap1 oxidative stress signal transduction pathway found in Saccharomyces cerevisiae is redox-regulated. We have examined the thermodynamic basis of the disulfide/dithiol couples that are involved in the regulation of this pathway. The oxidized form of the Yap1 redox domain (Yap1-RD) fragment, derived from the Yap1 transcription factor, contains two disulfide bonds, one between Cys303 and Cys598 and one between Cys310 and Cys629. Oxidation-reduction titrations reveal the presence of two separate two-electron redox couples in Yap1-RD, with redox midpoint potentials (E-m) of -155 and -330 mV, respectively, at pH 7.0. We measured Em values of -275 and -265 mV for the two cytoplasmic S. cerevisiae thioredoxins, Trx1 and Trx2, respectively, both at pH 7.0. Last, we measured an E-m value of -255 mV for the Cys36-Cys82 disulfide bond at pH 6.0 in the glutathione peroxidase-like enzyme, oxidant receptor protein (Orp1). We were unable to obtain satisfactory redox titration data for Orp1 at pH 7.0, but if the redox-active disulfide of Orp1 exhibits the -59 mV per pH unit dependence for E-m typical of protein disulfides in this pH region, an Em value of -315 mV can be estimated for Orp1 at pH 7.0 by extrapolation. Together, these data suggest that, at physiological ratios of Trx(OX)/Trx(red), the reduction of both the E-m = -315 mV disulfide of Orp1 and the E-m = -330 mV disulfide of Yap1 by either Trx1 or Trx2 would be thermodynamically possible.