Novel roles for Fe-S clusters in stabilizing or generating radical intermediates

The role and properties of the Fe-S clusters in spinach ferredoxin-thioredoxin reductase (FTR), E. coli biotin synthase and E. coli pyruvate formatelyase activating enzyme (PFL-AE) have been investigated by the combination of EPR, resonance Raman, and UV-visible absorption and variable temperature magnetic circular dichroism spectroscopies. FTR is shown to be a novel class of disulfide reductase with an active site involving a [4Fe-4S] cluster and an adjacent cysteine disulfide. The results suggest that the cluster stabilizes the one-electron reduced intermediate of the enzymatic reaction by forming a covalent adduct with one of the cysteines of the active site disulfide, leaving the other cysteine available to form the heterodisulfide adduct with the substrate disulfide. Biotin synthase and PFL-AE are both shown to be homodimeric enzymes with subunit-bridging [4Fe-4S] clusters that undergo novel [4Fe-4S]²⁺ ⇄ 2[2Fe-2S]²⁺ + 2e^- cluster conversions. It is proposed that the [4Fe-4S] clusters in both enzymes are directly involved with generating a 5′-deoxyadenosyl radical from S-adenosyl-L-methionine and that the oxidative cluster conversion provides a means of regulating enzyme activity on exposure to O₂ without irreversible cluster degradation. The possibility that the catalytic roles for the [4Fe-4S] clusters in both classes of enzyme investigated in this work involve novel μ₃-S^2--based cluster chemistry is discussed.