title: A HEAT-repeats containing protein, IaiH, stabilizes the iron-sulfur cluster bound to the cyanobacterial IscA homologue, IscA2.
abstract: IscA homologues are involved in iron-sulfur cluster biosynthesis. In the non-nitrogen-fixing cyanobacterium Synechocystis PCC 6803, there are two IscA homologues, SLR1417 and SLR1565 (designated IscA1 and IscA2), of which only IscA2 exists as a protein complex with the HEAT-repeat-containing protein, SLR1098 (IaiH). We observed that the absorption spectrum of the recombinant IscA2/IaiH complex resembles that of IscA2 alone, although it is sharper. In the presence of dithiothreitol, the [2Fe-2S] cluster of IscA2 alone, but not of the IscA2/IaiH complex, became reductively labile upon the addition of sodium dithionite. This implies that the IscA2 moiety of the [2Fe-2S] cluster is stabilized by the presence of IaiH. The [2Fe-2S] cluster of the IscA2/IaiH complex was destabilized by sodium dithionite in the absence of dithiothreitol, suggesting that the in vivo stability of the iron-sulfur cluster in the IscA2/IaiH complex is influenced by the redox state of cellular thiols. When any one of three conserved cysteine residues in IscA2, potential ligands for the [2Fe-2S] cluster, was replaced with serine, the amount of assembled [2Fe-2S] cluster and protein complex was significantly reduced in E. coli cells. The cysteine mutated IscA2/IaiH complexes that were present all contained a [2Fe-2S]-like cluster suggesting that the assembly of a stable iron-sulfur cluster bound to IscA2 is required for efficient and stable complex formation. Truncated IaiH proteins were analyzed using the yeast two-hybrid assay to identify the essential domain of IaiH that interacts physically with IscA2. At least 2 of the 5 N-terminal HEAT repeats of IaiH were found to be required for interaction with IscA2.
authors.affiliation: Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita 565-0871.
authors.forename: Kozo
authors.initials: K
authors.lastname: Morimoto
authors.forename: Shusei
authors.initials: S
authors.lastname: Sato
authors.forename: Satoshi
authors.initials: S
authors.lastname: Tabata
authors.forename: Masato
authors.initials: M
authors.lastname: Nakai
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000458:Cyanobacteria; D003545:Cysteine / Q000235:genetics; D004227:Dithionite / Q000737:chemistry; D004492:Edetic Acid / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid; D013056:Spectrophotometry, Ultraviolet; D020798:Two-Hybrid System Techniques
chemical_list: D001426:Bacterial Proteins; D007506:Iron-Sulfur Proteins; D011994:Recombinant Proteins; D004227:Dithionite; D004492:Edetic Acid; D003545:Cysteine