The CCG-domain-containing subunit SdhE of succinate:quinone oxidoreductase from Sulfolobus solfataricus P2 binds a [4Fe-4S] cluster.
In type E succinate:quinone reductase (SQR), subunit SdhE (formerly SdhC) is thought to function as monotopic membrane anchor of the enzyme. SdhE contains two copies of a cysteine-rich sequence motif (CX(n)CCGX(m)CXXC), designated as the CCG domain in the Pfam database and conserved in many proteins. On the basis of the spectroscopic characterization of heterologously produced SdhE from Sulfolobus tokodaii, the protein was proposed in a previous study to contain a labile [2Fe-2S] cluster ligated by cysteine residues of the CCG domains. Using UV/vis, electron paramagnetic resonance (EPR), (57)Fe electron-nuclear double resonance (ENDOR) and Mössbauer spectroscopies, we show that after an in vitro cluster reconstitution, SdhE from S. solfataricus P2 contains a [4Fe-4S] cluster in reduced (2+) and oxidized (3+) states. The reduced form of the [4Fe-4S](2+) cluster is diamagnetic. The individual iron sites of the reduced cluster are noticeably heterogeneous and show partial valence localization, which is particularly strong for one unique ferrous site. In contrast, the paramagnetic form of the cluster exhibits a characteristic rhombic EPR signal with g (zyx) = 2.015, 2.008, and 1.947. This EPR signal is reminiscent of a signal observed previously in intact SQR from S. tokodaii with g (zyx) = 2.016, 2.00, and 1.957. In addition, zinc K-edge X-ray absorption spectroscopy indicated the presence of an isolated zinc site with an S(3)(O/N)(1) coordination in reconstituted SdhE. Since cysteine residues in SdhE are restricted to the two CCG domains, we conclude that these domains provide the ligands to both the iron-sulfur cluster and the zinc site.
Hamann
Nils
N
Max Planck Institute for Terrestrial Microbiology, Karl-von-Frisch-Strasse, 35043, Marburg, Germany.
Bill
Eckhard
E
Shokes
Jacob E
JE
Scott
Robert A
RA
Bennati
Marina
M
Hedderich
Reiner
R
D020816:Amino Acid Motifs; D001665:Binding Sites; D004578:Electron Spin Resonance Spectroscopy; D042963:Electron Transport Complex II / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics; D013056:Spectrophotometry, Ultraviolet; D015204:Spectroscopy, Mossbauer; D048229:Sulfolobus solfataricus / Q000201:enzymology*; D015032:Zinc / Q000737:chemistry
D007506:Iron-Sulfur Proteins; D021122:Protein Subunits; D042963:Electron Transport Complex II; D015032:Zinc