title: An Isc-type extremely thermostable [2Fe-2S] ferredoxin from Aquifex aeolicus. Biochemical, spectroscopic, and unfolding studies.
abstract: Analysis of the genome of the hyperthermophilic bacterium Aquifex aeolicus has revealed the presence of a previously undetected gene potentially encoding a plant- and mammalian-type [2Fe-2S] ferredoxin. Expression of that gene in Escherichia coli has yielded a novel thermostable [2Fe-2S] ferredoxin (designated ferredoxin 5) whose sequence is most similar to those of ferredoxins involved in the assembly of iron-sulfur clusters (Isc-Fd). It nevertheless differs from the latter proteins by having deletions near its N- and C-termini, and no cysteine residues other than those involved in [2Fe-2S] cluster coordination. Resonance Raman, low-temperature MCD and EPR studies show close spectral similarities between ferredoxin 5 and the Isc-Fd from Azotobacter vinelandii. Mössbauer spectra of the reduced protein were analyzed with an S = 1/2 spin Hamiltonian and interpreted in the framework of the ligand field model proposed by Bertrand and Gayda. The redox potential of A. aeolicus ferredoxin 5 (-390 mV) is in keeping with its relatedness to Isc-Fd. Unfolding experiments showed that A. aeolicus ferredoxin 5 is highly thermostable (T(m) = 106 degrees C at pH 7), despite being devoid of features (e.g., high content of charged residues) usually associated with extreme thermal stability. Searches for genes potentially encoding plant-type [2Fe-2S] ferredoxins have been performed on the sequenced genomes of hyperthermophilic organisms. None other than the two proteins from A. aeolicus were retrieved, indicating that this otherwise widely distributed group of proteins is barely represented among hyperthermophiles.
authors.affiliation: Département de Réponse et Dynamique Cellulaires, CEA-Grenoble, 38054 Grenoble, France.
authors.forename: Géraldine
authors.initials: G
authors.lastname: Mitou
authors.forename: Catherine
authors.initials: C
authors.lastname: Higgins
authors.forename: Pernilla
authors.initials: P
authors.lastname: Wittung-Stafshede
authors.forename: Richard C
authors.initials: RC
authors.lastname: Conover
authors.forename: Archer D
authors.initials: AD
authors.lastname: Smith
authors.forename: Michael K
authors.initials: MK
authors.lastname: Johnson
authors.forename: Jacques
authors.initials: J
authors.lastname: Gaillard
authors.forename: Audria
authors.initials: A
authors.lastname: Stubna
authors.forename: Eckard
authors.initials: E
authors.lastname: Münck
authors.forename: Jacques
authors.initials: J
authors.lastname: Meyer
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004578:Electron Spin Resonance Spectroscopy; D005288:Ferredoxins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D006358:Hot Temperature; D007506:Iron-Sulfur Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D011489:Protein Denaturation; D017510:Protein Folding; D013056:Spectrophotometry, Ultraviolet; D015204:Spectroscopy, Mossbauer; D013059:Spectrum Analysis, Raman
chemical_list: C081714:2Fe-2S ferredoxin; D005288:Ferredoxins; D007506:Iron-Sulfur Proteins