title: Characterization of a mutated rubredoxin with a cysteine ligand of the iron replaced by serine.
abstract: The active site of rubredoxins consists of a single iron tetrahedrally coordinated to four cysteinate sulfurs. One of the iron ligands, cysteine 42, has been mutated into serine in Clostridium pasteurianum rubredoxin. This mutation resulted in a shift to higher energy of the 320-800 nm region of the UV-visible absorption spectrum. Resonance Raman spectra showed that the nu 1 breathing mode of the iron chromophore was upshifted as a result of the C42S mutation. The spectral pattern, however, was not largely disturbed by the mutation. The EPR spectra of both the wild type and the C42S mutated protein displayed the characteristic features, at g = 4.3 and g = 9.5, of the "3/2" and "1/2" Kramers' doublets, respectively, of a S = 5/2 multiplet. These combined data afford strong evidence that in the C42S mutated rubredoxin serine has replaced cysteine 42 as a ligand of the iron, while maintaining the tetrahedral coordination of the metal. The most spectacular effect of the C42S mutation was a ca. 200 mV downshift of the redox potential of rubredoxin.
authors.affiliation: CEA-Département de Biologie Moléculaire et Structurale, CENG, Grenoble, France.
authors.forename: J
authors.initials: J
authors.lastname: Meyer
authors.forename: J
authors.initials: J
authors.lastname: Gaillard
authors.forename: M
authors.initials: M
authors.lastname: Lutz
mesh_terms: D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D003013:Clostridium / Q000235:genetics; D003545:Cysteine / Q000737:chemistry / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004578:Electron Spin Resonance Spectroscopy; D007501:Iron / Q000737:chemistry; D008024:Ligands; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D012416:Rubredoxins / Q000737:chemistry / Q000235:genetics*; D012694:Serine / Q000737:chemistry / Q000235:genetics; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet; D013059:Spectrum Analysis, Raman
chemical_list: D017931:DNA Primers; D008024:Ligands; D012416:Rubredoxins; D012694:Serine; D007501:Iron; D003545:Cysteine