Abstract
In plants, the cysteine desulfurase (AtNFS1) and frataxin (AtFH) are involved in the formation of Fe-S groups in mitochondria, specifically, in Fe and sulfur loading onto scaffold proteins, and the subsequent formation of the mature Fe-S cluster. We found that the small mitochondrial chaperone, AtISD11, and AtFH are positive regulators for AtNFS1 activity in Arabidopsis. Moreover, when the three proteins were incubated together, a stronger attenuation of the Fenton reaction was observed compared to that observed with AtFH alone. Using pull-down assays, we found that these three proteins physically interact, and sequence alignment and docking studies showed that several amino acid residues reported as critical for the interaction of their human homologous are conserved. Our results suggest that AtFH, AtNFS1 and AtISD11 form a multiprotein complex that could be involved in different stages of the iron–sulfur cluster (ISC) pathway in plant mitochondria.
Highlights
IntroductionFour different types of Fe-S cluster biosynthetic systems have been described: (1) the NIF (nitrogen fixation) system, present in azeotropic bacteria and required for metallocluster biogenesis in nitrogenase [3,4]; (2) the SUF (sulfur mobilization) system, originally identified in E. coli and found in plastids; (3) the ISC (iron–sulfur cluster) system, commonly found in bacteria and mitochondria [5,6]; and (4) the CIA (cytosolic iron-sulfur cluster assembly) system, for cytosolic/nuclear proteins from eukaryotic cells [7]
In Arabidopsis, there are two genes that code for cysteine desulfurases involved in the synthesis of Fe-S groups: AtNFS1, which is located in mitochondria, and AtNFS2, located in chloroplasts [8,14,15,16,17,18]
In the presence of AtFH and AtISD11 (1:1:1 molar ratio), the desulfurase activity showed an increment of about 6-fold in the V max (47.83 ± 3.52 U/mg) and an increase of about 3.5-fold in the apparent affinity for cysteine (S0.5 = 0.23 ± 0.04 mM) respect to the assay using AtNFS1 alone (Figure 1). These results indicate that both, AtISD11 and AtFH would acts as regulators, increasing the desulfurase activity, suggesting that the three proteins could interact forming a multiprotein complex
Summary
Four different types of Fe-S cluster biosynthetic systems have been described: (1) the NIF (nitrogen fixation) system, present in azeotropic bacteria and required for metallocluster biogenesis in nitrogenase [3,4]; (2) the SUF (sulfur mobilization) system, originally identified in E. coli and found in plastids; (3) the ISC (iron–sulfur cluster) system, commonly found in bacteria and mitochondria [5,6]; and (4) the CIA (cytosolic iron-sulfur cluster assembly) system, for cytosolic/nuclear proteins from eukaryotic cells [7]. In Arabidopsis, there are two genes that code for cysteine desulfurases involved in the synthesis of Fe-S groups: AtNFS1, which is located in mitochondria, and AtNFS2 (cpNifS), located in chloroplasts [8,14,15,16,17,18]
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