Iron–sulfur proteins are proteins characterized by the presence of iron–sulfur clusters containing sulfide-linked di-, tri-, and tetrairon centers in variable oxidation states. Iron–sulfur clusters are found in a variety of metalloproteins, such as the ferredoxins, as well as NADH dehydrogenase, hydrogenases, coenzyme Q – cytochrome c reductase, succinate – coenzyme Q reductase and nitrogenase. Iron–sulfur clusters are best known for their role in the oxidation-reduction reactions of electron transport in mitochondria and chloroplasts. Both Complex I and Complex II of oxidative phosphorylation have multiple Fe–S clusters. They have many other functions including catalysis as illustrated by aconitase, generation of radicals as illustrated by SAM-dependent enzymes, and as sulfur donors in the biosynthesis of lipoic acid and biotin. Additionally, some Fe–S proteins regulate gene expression. Fe–S proteins are vulnerable to attack by biogenic nitric oxide, forming dinitrosyl iron complexes. In most Fe–S proteins, the terminal ligands on Fe are thiolate, but exceptions exist. The prevalence of these proteins on the metabolic pathways of most organisms leads some scientists to theorize that iron–sulfur compounds had a significant role in the origin of life in the iron–sulfur world theory. Structural motifs In almost all Fe–S proteins, the Fe centers are tetrahedral and the terminal ligands are thiolato sulfur centers from cysteinyl residues. The sulfide groups are either two- or three-coordinated. Three distinct kinds of Fe–S clusters with these features are most common. Structure-Function Principles To serve their various biological roles, iron-sulfur proteins effect rapid electron transfers and span the whole range of physiological redox potentials from -600 mV to +460 mV. Iron-sulfur proteins are involved in various biological electron transport processes, such as photosynthesis and cellular respiration, which require rapid electron transfer to sustain the energy or biochemical needs of the organism. Fe3+-SR bonds have unusually high covalency which is expected. When comparing the covalency of Fe3+ with the covalency of Fe2+, Fe3+ has almost double the covalency of Fe2+ (20% to 38.4%). Fe3+ is also much more stabilized than Fe2+. Hard ions like Fe3+ normally have low covalency because of the energy mismatch of the metal Lowest Unoccupied Molecular Orbital with the ligand Highest Occupied Molecular Orbital. There is HO-H—S-Cys H-bonding from external H2O’s positioned by the protein close to the active site and this H-bonding decreases the lone pair electron donation from the Cys-S donor to the Fe3+/2+. Using lyophilization to remove these external H2O’s results in increased Fe-S covalency, which