Electron Bifurcation Arises from Emergent Features of Multicofactor Enzymes

Quinone-based electron bifurcation (QBEB) catalyzed by cytochrome bc ₁ (cyt. bc ₁) plays a critical role in maximizing the efficiency of biological energy conversion. The canonical QBEB model, grounded in equilibrium redox potentials, dictates the order of QBEB steps with initial endergonic reduction of a high-potential iron-sulfur cluster (2Fe2S) by quinol followed by exergonic reduction of low-potential heme b _L by semiquinone (SQ). However, this concept falls short in explaining several experimental observations, including intermediate semiquinone spin-coupled to 2Fe2S (SQ-2Fe2S^red) and the absence of short-circuiting. The current DFT calculations, performed on large cluster models of cyt. bc ₁, encompassing both 2Fe2S and heme b _L, identified the location of donor (HOMO) and acceptor (LUMO) orbitals along with the previously unconsidered microstates to reveal that QBEB is an emergent property of an integrated system of redox cofactors where transient charge separations dynamically modulate electron affinities. In this system, electron transfer initiates preferentially toward heme b _L, indicating a departure from the conventional sequence proposed by the canonical QBEB. Based on this finding, we introduce an emergent electron transfer (EMET) model of QBEB and demonstrate that its assumptions are supported by electron paramagnetic resonance spectroscopy data. Unlike the canonical QBEB, EMET proposes a relatively flat energy profile for the QBEB that accommodates stable SQ-2Fe2S^red and explains suppression of short circuits without additional assumptions. It highlights the importance of state-dependent electrostatic interactions in shaping electron transfer pathways. In general, the concept of emergence inherent to EMET offers a mechanistic framework applicable to a broad range of multicofactor redox enzymes beyond cyt. bc ₁.