The Apparently Unreactive Substrate Facilitates the Electron Transfer for Dioxygen Activation in Rieske Dioxygenases

Rieske dioxygenases belong to the non-heme iron family of oxygenases and catalyze important cis-dihydroxylation as well as O-/N-dealkylation and oxidative cyclization reactions for a wide range of substrates. The lack of substrate coordination at the non-heme ferrous iron center, however, makes it particularly challenging to delineate the role of the substrate for productive O2 activation. Here, we studied the role of the substrate in the key elementary reaction leading to O2 activation from a theoretical perspective by systematically considering (i) the 6-coordinate to 5-coordinate conversion of the non-heme Fe-II upon abstraction of a water ligand, (ii) binding of O2 , and (iii) transfer of an electron from the Rieske cluster. We systematically evaluated the spin-state-dependent reaction energies and structural effects at the active site for all combinations of the three elementary processes in the presence and absence of substrate using naphthalene dioxygenase as a prototypical Rieske dioxygenase. We find that reaction energies for the generation of a coordination vacancy at the non-heme FeII center through thermoneutral H2O reorientation and exothermic O2 binding prior to Rieske cluster oxidation are largely insensitive to the presence of naphthalene and do not lead to formation of any of the known reactive Fe-oxygen species. By contrast, the role of the substrate becomes evident after Rieske cluster oxidation and exclusively for the 6-coordinate non-heme FeII sites in that the additional electron is found at the substrate instead of at the iron and oxygen atoms. Our results imply an allosteric control of the substrate on Rieske dioxygenase reactivity to happen prior to changes at the non-heme FeII in agreement with a strategy that avoids unproductive O2 activation.

Automated summary

In this study, the role of the substrate in the O2 activation process of Rieske dioxygenase was investigated theoretically. The results showed that the presence of substrate did not affect the reaction energies and structural effects at the non-heme FeII center, but played a role in regulating the reactivity of Rieske dioxygenase.