Mechanism of Mixed-Valence Fe(2.5+)···Fe(2.5+) Formation in Fe(4)S(4) Clusters in the Ferredoxin Binding Motif

[Image: see text] Most low-potential Fe(4)S(4) clusters exist in the conserved binding sequence CxxCxxC (C(n)C(n+3)C(n+6)). Fe(II) and Fe(III) at the first (C(n)) and third (C(n+6)) cysteine ligand sites form a mixed-valence Fe(2.5+)···Fe(2.5+) pair in the reduced Fe(II)(3)Fe(III) cluster. Here, we investigate the mechanism of how the conserved protein environment induces mixed-valence pair formation in the Fe(4)S(4) clusters, F(X), F(A), and F(B) in photosystem I, using a quantum mechanical/molecular mechanical approach. Exchange coupling between Fe sites is predominantly determined by the shape of the Fe(4)S(4) cluster, which is stabilized by the preorganized protein electrostatic environment. The backbone NH and CO groups in the conserved CxxCxxC and adjacent helix regions orient along the Fe(Cn)···Fe(C(n+6)) axis, generating an electric field and stabilizing the Fe(Cn)(II)Fe(C(n+6))(III) state in F(A) and F(B). The overlap of the d orbitals via −S– (superexchange) is observed for the single Fe(Cn)(II)···Fe(C(n+6))(III) pair, leading to the formation of the mixed-valence Fe(2.5+)···Fe(2.5+) pair. In contrast, several superexchange Fe(II)···Fe(III) pairs are observed in F(X) due to the highly symmetric pair of the CDGPGRGGTC sequences. This is likely the origin of F(X) serving as an electron acceptor in the two electron transfer branches.