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bioRxiv · 10.64898/2026.01.14.699433

Ubiquinone is a hysteretic modulator of the NADH:cytochrome b5 reductase activity of human Cb5R

Valerio, G. N.·Martinez-Costa, O. H.·Sanchez-Cabeza, C.·Cordas, C. M.·Samhan-Arias, A. K.
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Abstract

BackgroundCytochrome b5 reductase is a flavoprotein that transfers electrons from NADH to multiple electron acceptors, such as cytochrome b5 or ubiquinone. Hysteresis is a phenomenon characterized by a slow transition between active and inactive catalytic states, leading to a lag phase in enzymatic activity. In this study, the effect of the soluble analogue of ubiquinone named 2,3 dimethoxy-5-methyl-1,4 benzoquinone (CoQ0) on the NADH:Cb5 reductase activity of recombinant human soluble Cb5R, using recombinant human soluble Cb5 as a substrate was evaluated. The aim of this study was to determine whether ubiquinone exerts a hysteretic modulation of this activity based on previous studies supporting that microsomal reduction of cytochrome b5 is controlled by redox hysteresis. ResultsThe NADH:cytochrome b5 reductase activity of Cb5R was characterized at different concentrations of Cb5R, cytochrome b5, and CoQ0 by monitoring the reduction of cytochrome b5. The addition of CoQ0 induced the appearance of a lag phase, whose duration increased with the concentration of CoQ0 and decreased with higher concentrations of cytochrome b5 or Cb5R. Additionally, a concentration-dependent decrease in the maximum rate of reduction and the appearance of positive cooperativity was observed in the presence of CoQ0 which resulted in leading to lower KM values for cytochrome b5. This suggests the formation of a CoQ0:Cb5R complex altering the interaction between the reductase and cytochrome b5 which increases the affinity for cytochrome b5. Cyclic voltammetry data support the formation of CoQ0/protein complex that could be responsible for the hysteretic behavior. ConclusionsThese results support the hypothesis that CoQ0 is a hysteretic modulator and inhibitor of the NADH: cytochrome b5 reductase activity of human Cb5R.

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Valerio, G. N., Martinez-Costa, O. H., Sanchez-Cabeza, C., Cordas, C. M., Samhan-Arias, A. K.. 2026-01-14. Ubiquinone is a hysteretic modulator of the NADH:cytochrome b5 reductase activity of human Cb5R. https://doi.org/10.64898/2026.01.14.699433

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