Heterogeneous subunit structures in the pyranose 2-oxidase homotetramer revealed by theoretical analysis of the rates of photoinduced electron transfer from a tryptophan to the excited flavin

•The extraordinary heterogeneous fluorescence decays of pyranose 2-oxidase were systematically analyzed with atomic coordinates obtained by a molecular dynamics simulation and the ET rate calculation by Kakitani and Mataga ET theory.•All possible combinations of the four subunits revealed that the slow component was from subunit A and the fast component was from the other three subunits.•The large difference in the fast and slow fluorescent lifetimes is ascribed to the difference in the standard free energy gap related to electron affinity of Iso*.

Pyranose 2-oxidase (P2O) from Trametes multicolor forms a homotetramer in which each of the subunits contains flavin adenine dinucleotide (FAD). The fluorescence of P2O decays with two lifetime components; a slow (358 ps) and a fast (∼90 fs) decay. The lifetime of the fast component is emission-wavelength dependent and is ascribed to fast photoinduced electron transfer (ET) from Trp168 to the excited isoalloxazine (Iso*) in FAD. The donor–acceptor distances were ∼0.7 nm. The extraordinary heterogeneous decays were analyzed with atomic coordinates obtained by a molecular dynamics simulation and the ET rate by Kakitani and Mataga. The emission-wavelength dependent decays in the fast component were elucidated by introducing emission-wavelength dependent standard free energy related to electron affinity of Iso*. Examination of all possible combinations of the four subunits revealed that the slow component was from subunit A and the fast component was from the other three subunits. Agreements between the observed and calculated decays were all excellent. The large difference in the fast and slow fluorescent lifetimes is ascribed to the difference in the standard free energy gap related to electron affinity of Iso*. The dependence of the logarithmic ET rates on the center-to-center distance displayed approximate linear functions (Dutton rule) when the rate was relatively slow and parabolic functions when the rate was ultrafast. The Dutton rule originated from the exponential term of the ET rate, not from the electronic coupling term.