Alternative ground states enable pathway switching in biological electron transfer

Abstract

Electron transfer is the simplest chemical reaction and constitutes the basis of a large variety of biological processes, such as photosynthesis and cellular respiration. Nature has evolved specific proteins and cofactors for these functions. The mechanisms optimizing biological electron transfer have been matter of intense debate, such as the role of the protein milieu between donor and acceptor sites. Here we propose a mechanism regulating long-range electron transfer in proteins. Specifically, we report a spectroscopic, electrochemical, and theoretical study on WT and single-mutant Cu(A) redox centers from Thermus thermophilus, which shows that thermal fluctuations may populate two alternative ground-state electronic wave functions optimized for electron entry and exit, respectively, through two different and nearly perpendicular pathways. These findings suggest a unique role for alternative or "invisible" electronic ground states in directional electron transfer. Moreover, it is shown that this energy gap and, therefore, the equilibrium between ground states can be fine-tuned by minor perturbations, suggesting alternative ways through which protein-protein interactions and membrane potential may optimize and regulate electron-proton energy transduction.

Fig. 1.

(A) The Cu_A site from Thermus thermophilus ba₃ oxidase (rendered from PDB ID 2CUA) (35). (B) Double-well potential describing the ground-state symmetry as a function of the Cu—Cu distance, including the calculated σ*_u (blue) and π_u (red) molecular orbitals. The large splitting between both states yields an adiabatic surface along the coordinate.

Fig. 2.

(A) ¹³C NMR spectra highlighting resonances from the cysteine residues in WT Cu_A and the M160H and M160Q mutants. (B) Experimental absorption spectra for WT Cu_A and the M160H and M160Q mutants. (C) Simulated spectra for σ*_u and π_u ground states on the WT geometry.

Fig. 3.

(Upper) Optimal ET pathway (red) from cytochrome c to Cu_A in the π_u state, and (Lower) from Cu_A in the σ_u* state to the ba₃ group of the oxidase. Rendered from the structure of T. thermophilus ba₃ oxidase (PDB ID code 1XME) (41) and the model complex between the Cu_A domain and cytochrome c₅₅₂ (PDB ID code 2FWL) (13).