STRUCTURAL STUDIES ON METAL SUBSTITUTED DERIVATIVES OF DESULFOREDOXIN, A RUBREDOXIN TYPE PROTEIN
M. Archer1, A.L. Carvalho1, S. Teixeira1, I. Moura2, J.J.G Moura2, M.J. Romao1
1- I.T.Q.B., Apt. 127, 2780 Oeiras, Portugal
and, I.S.T., Dep. Química, 1096-Lisboa Codex
2- F.C.T- U.N.L., Dep. Química, 2825 Monte Caparica, Portugal
E-mail: archer@itqb.unl.pt
Keywords: Fe(Cys)4 tetrahedral
coordination, metal substitution, rubredoxin type protein
The crystal structure of desulforedoxin from Desulfovibrio gigas, a new homo-dimeric (2x36 aminoacids) non-heme iron protein, has been solved at 1.8 Å resolution by the SIRAS method using the indium substituted protein as the single derivative [1]. The two-fold symmetric dimer is firmly hydrogen bonded and folds as an incomplete b-barrel with both iron centers placed on opposite poles of the molecule. Each iron atom is coordinated to four cysteinyl residues in a distorted tetrahedral arrangement. Desulforedoxin and rubredoxin backbones share some structural features but show significant differences in terms of metal environment and water structure, which account for the known spectroscopic differences between rubredoxin and desulforedoxin [2].
The simplicity of the active center in Dx and the possibility of reconstituting the apoprotein make this protein suitable for the crystallographic analysis of metal-substituted derivatives. This study is important to obtain model compounds that can mimic the tetrahedral sulfur coordination of metals in biology.
Several metal replacement experiments have been successfully carried out with Co, Zn, Mn, V, Ga, Cd, Hg and Ni. These metal reconstituted Dx crystallize under different experimental conditions yielding several crystal morphologies. The Ni substituted Dx structure is of particular interest, as this protein and Ni substituted rubredoxins were shown to mimic the reactivity pattern of Ni-containing hydrogenases with respect to hydrogen production, deuterium-proton and inhibition by carbon monoxide [3]. Recently, the structures of Dx substituted with Ga, In, Cd and Hg have been determined by the molecular replacement method and refined.
The comparison of the metal derivatives three-dimensional
structures shows that the overall secondary and tertiary
structure of the metal derivatives is maintained, while some
differences in metal coordination geometry occur, namely bond
lengths and angles of the metal with the sulfur ligands. The
effects of changing d-shell electrons in the protein structure
and correlation with spectroscopic and redox properties of the
metal center geometry are also possible.