EPR and Electron Spin Echo Studies of Iron-Sulfur Clusters S-1 And S-2 in Bovine Heart Succinate Dehydrogenase

Abstract

The ligands of the three iron-sulfur clusters in bovine heart succinate dehydrogenase (BH-SDH) have not been determined to date. We have made a detailed comparison of the electron spin echo envelope modulation (ESEEM) spectra from the [2Fe-2S] (+1,+2) cluster S-I with spectra obtained from two better-characterized iron-sulfur proteins. X- ray crystallographic data (1) demonstrated that the [2Fe-2S](+1,+2) cluster in Spirulina platensis ferredoxin has four cysteine sulfur ligands, but that there are several nitrogens in the polypeptide backbone that are close enough to form NH..S hydrogen bonds to the cluster. The ESEEM spectrum of the dithionite-reduced protein shows intense modulations characteristic of 14N at all g-values. At g = 1.94, four distinct modulation frequencies are evident: 0.6, 1.9, 2.9 and 4.4 MHz. The ESEEM spectrum of spinach ferredoxin is virtually identical to that of S. platensis feredoxin at g = 1.94, and at the other principal g-values. The ESEEM spectrum of cluster S-I in BH-SDH contains four very similar frequencies at g = 1.94. It does not exhibit any additional modulation frequencies which might be attributed to nitrogen from an imidazole ligand. The data indicate that cluster S-I also has only non- imidazole ligands, probably all sulfur.