ASSEMBLY OF A [2FE-2S](2-PASTEURIANUM RUBREDOXIN() CLUSTER IN A MOLECULAR VARIANT OF CLOSTRIDIUM)

Citation
J. Meyer et al., ASSEMBLY OF A [2FE-2S](2-PASTEURIANUM RUBREDOXIN() CLUSTER IN A MOLECULAR VARIANT OF CLOSTRIDIUM), Biochemistry, 36(43), 1997, pp. 13374-13380
Citations number
46
Categorie Soggetti
Biology
Journal title
ISSN journal
00062960
Volume
36
Issue
43
Year of publication
1997
Pages
13374 - 13380
Database
ISI
SICI code
0006-2960(1997)36:43<13374:AOA[RC>2.0.ZU;2-8
Abstract
The rubredoxin from Clostridium pasteurianum contains a single iron at om bound to the polypeptide chain by cysteines 6, 9, 39, and 42. The C 42A variant of this protein has been prepared by site-directed mutagen esis and heterologous expression of the gene in Escherichia coli, The mutated protein was found to contain an unexpected chromophore that ha s been characterized by a variety of techniques, UV-visible absorption and resonance Raman spectra were strongly reminiscent of those of [2F e-2S] proteins, Mossbauer spectra of the oxidized chromophore isolated in oxygen-free conditions indicated low-temperature diamagnetism resu lting from antiferromagnetically coupled high-spin ferric ions. Analys is of X-ray absorption fine structure spectra yielded an Fe-Fe distanc e of 2.68 Angstrom, Colorimetric assays of iron and inorganic sulfide showed that the two elements are present in a 1:1 ratio, Electrospray- ionization mass spectra displayed a major component at M = 6190 Da, i. e. the molecular mass of the C42A apoprotein plus two atomic masses of iron and two atomic masses of sulfur. Taken together, these data show that a mere point mutation allows the stabilization of a binuclear [2 Fe-2S] cluster in a protein that normally accommodates a mononuclear F e(Scys)(4) site, Assembly of a [2Fe-2S] cluster may occur because rubr edoxin assumes a similar fold around its metal center as the [2Fe-2S] Rieske protein, Alternatively, a more extensive structural rearrangeme nt of the polypeptide chain of the C42A rubredoxin variant may be cons idered as well.