PARENT SESSION
Posters P2A Type I reaction centres. Abstracts (181-218)

Iron sulfur cluster assembly in Photosystem I: The role of the transcriptional repressor SufR in regulating the suf regulon in cyanobacterium Synechococcus sp. PCC 7002. Ramakrishnan Balasubramanian^*,1, Tao Wang¹, Gaozhong Shen¹, Lee Hoffart¹, Carsten Krebs^{1, 2}, Lee McIntosh³, Donald Bryant¹, John Golbeck^{1, 2}, ¹ Department of Biochemistry and Molecular Biology, University Park, PA, United States² Department of Chemistry, University Park, PA, United States³ MSU-DOE Plant Research Laboratory, East Lansing, MI, United States

ABSTRACT- The suf regulon in Synechocystis sp. PCC 6803 consists of the sufA, sufB, sufC, sufD, sufE and sufS genes. The ORF sll0088 is located just upstream of the sufBCDS operon and is involved in regulating the Fe/S cluster assembly in Photosystem I (Yu et al., (2003) J. Bact. 185, 3878-3887). Northern blot and RT-PCR analyses confirm the prediction that the sll0088 gene product serves as the transcriptional repressor of the suf regulon. The protein encoded by sll0088 has two significant features: (i) a helix-loop-helix DNA-binding domain near the N terminus, and (ii) four highly conserved cysteine residues near the C terminus that may bind a Fe/S cluster. Sll0088 is therefore renamed sufR. The SufR protein was overexpressed in Escherichia coli and purified to homogeneity. EPR and Mössbauer spectroscopy of the Fe/S reconstituted SufR protein show the presence of a [4Fe-4S] cluster. Size exclusion chromatography together with iron and labile sulfide analyses indicates that SufR is a homodimer that co-ordinates one [4Fe-4S] cluster. Site directed mutagenesis shows that the third cysteine in SufR is not involved in ligating the Fe/S cluster. Because only two of the four cysteines in SufR likely co-ordinate the Fe/S cluster, the other cysteines may have a structural or regulatory role. Our experiments establish the role of SufR as a transcriptional repressor for the suf genes, with its [4Fe-4S] cluster acting as a sensor. We suggest that when Fe/S clusters are abundant, SufR binds a cluster and the transcription of the suf genes is repressed; when Fe/S clusters are required, SufR does not bind a cluster and transcription of the suf genes is activated. The Fe/S cluster in SufR is oxygen sensitive, thereby limiting its lifetime. Fe/S cluster homeostatis is thus maintained in the cyanobacterial cell by a balance between synthesis and breakdown. Funded by USDA (to JHG).

KEY WORDS: SufR, [4Fe-4S], suf operon, Fe/S cluster assembly