Mössbauer and EPR studies of desulforedoxin from Desulfovibrio gigas.

Abstract

We have studied desulforedoxin, a nonheme iron protein isolated from Desulfovibrio gigas with Mossbauer and EPR spectroscopy.The protein, a dimer (Mr 7900) consisting of two identical subunits, contains two iron atoms in environments nondistinguishable by Mossbauer and EPR spectroscopy.Earlier sequence studies have established that each monomeric unit contains four cysteines in an unusual sequence.Studies on the sulfhydryl content of apo-and holoprotein suggest that all four cysteines are coordinated to the iron.The zero field splitting parameters D = (2.2 k 0.3) cm" and E / D = 0.08 of ferric desulforedoxin suggest geometrical differences between this protein and rubredoxin.T h e saturation field of -385 kG (corresponding to a hyperfine coupling constant A. = -21 MHz) indicates more covalent bonding than observed for rubredoxin (-410 kG).We have also obtained a set of fine and hyperfine structure parameters for reduced high spin ferrous desulforedoxin.Most interestingly the data show that the orbital ground state has predominantly dS-9 character while reduced rubredoxin has a ground state of 6 symmetry.Assuming that no other ligands but cysteine coordinate to the iron the data suggest that the electronic structure of the desulforedoxin chromophore is influenced by stereochemical constraints of the protein moiety.This study shows that protein aggregation, readily detected in the Mossbauer spectrum, can lead to a sizable underestimation of spin concentration from EPR experiments.We have also studied rubredoxin from D. gigas.The data are practically identical with those reported for the protein from Clostridium pasteurianum.Desulforedoxin, isolated from the sulfate-reducing bacterium Desulfouibrio gigas, is a small nonheme iron protein with some interesting spectral properties (1-3).The protein is isolated as a dimer ( M , = 7900) consisting of two identical subunits (2), with one iron atom and four cysteines per monomer.The visible spectrum, in its general features, resembles that of rubredoxin.Of special interest is the spacing of the