The chloroplast NifS-like protein of Arabidopsis thaliana is required for iron-sulfur cluster formation in ferredoxin.
Plastids are known to be able to synthesize their own iron-sulfur clusters, but the biochemical machinery responsible for this process is not known. In this study it is investigated whether CpNifS, the chloroplastic NifS-like cysteine desulfurase of Arabidopsis thaliana (L.) Heynh. is responsible for the release of sulfur from cysteine for the biogenesis of iron-sulfur (Fe-S) clusters in chloroplasts. Using an in vitro reconstitution assay it was found that purified CpNifS was sufficient for Fe-S cluster formation in ferredoxin in the presence of cysteine and a ferrous iron salt. Antibody-depletion experiments using stromal extract showed that CpNifS is also essential for the Fe-S cluster formation activity of chloroplast stroma. The activity of CpNifS in the stroma was 50- to 80-fold higher than that of purified CpNifS on a per-protein basis, indicating that other stromal factors cooperate in Fe-S cluster formation. When stromal extract was separated on a gel-filtration column, most of the CpNifS eluted as a dimer of 86 kDa, but a minor fraction of the stromal CpNifS eluted at a molecular weight of approx. 600 kDa, suggesting the presence of a multi-protein complex. The possible nature of the interacting proteins is discussed.
Ye
Hong
H
Department of Biology, Colorado State University, Anatomy/Zoology Building, Fort Collins, CO 80523, USA.
Garifullina
Gulnara F
GF
Abdel-Ghany
Salah E
SE
Zhang
Lihong
L
Pilon-Smits
Elizabeth A H
EA
Pilon
Marinus
M
D017360:Arabidopsis / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000378:metabolism*; D002736:Chloroplasts / Q000378:metabolism*; D002850:Chromatography, Gel; D005288:Ferredoxins / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000378:metabolism*; D007700:Kinetics
D029681:Arabidopsis Proteins; C472234:At1g08490 protein, Arabidopsis; D005288:Ferredoxins; D007506:Iron-Sulfur Proteins