Redox properties of protein disulfide bond in oxidized thioredoxin and lysozyme: A pulse radiolysis study

We have studied the one-electron reduction of oxidized Chlamydomonas reinha rdtii thioredoxin and compared it to that of hen egg white lysozyme, using CO2.- free radicals as reductants. This comparison shows that the thioredox in disulfide/thiol redox couple has different properties than that of lysoz yme: the disulfide radical pK(a) is much lower (around 5 for small disulfid es, 4.62 for lysozyme, <3 for thioredoxin). To get a better understanding o f the modulation of the thioredoxin redox properties we have constructed th e mutants W35A and D30A. Their reduction by pulse radiolysis indicates that W35 strongly controls both the disulfide radical acidity (the pK(a) in W35 A is equal to ca. 4), and the thiol reactivity. Asp30 is also involved in t he control of proton transfer to the disulfide free radical. In addition, i ts removal seems to increase the reduction potential of the thioredoxin thi yl/thiol couple. Overall, the reduction properties of thioredoxin confirm i ts nature as a unique reductant.