In this study, the effect of ubiquinone (CoQ 10 ) and its soluble analogue (CoQ 0 ) was measured on the NADH:C b 5 reductase activity of recombinant human membrane and soluble C b 5 R, respectively using soluble C b 5 as a substrate. The aim of this study was to determine whether quinones are inhibitors and can induce a hysteretic modulation of the activity, similar to that described for C b 5 reduction in microsomal systems. CoQ 0 and CoQ 10 inhibit the NADH:cytochrome b 5 reductase activity of soluble and membrane C b 5 R respectively, using soluble C b 5 as an electron acceptor. However, only CoQ 0 induced hysteresis of the soluble C b 5 R isoform. The addition of CoQ 0 induced the appearance of a lag phase, whose duration increased with the concentration of CoQ 0 and decreased with higher concentrations of soluble C b 5 or C b 5 R. Additionally, a concentration-dependent decrease in the maximum rate of reduction and the appearance of positive cooperativity was observed in the presence of CoQ 0 which resulted in lower K M values for C b 5 . This suggests the formation of a CoQ 0 :C b 5 R complex altering the interaction between the reductase and C b 5 which increases the affinity for C b 5 . Cyclic voltammetry data support the formation of CoQ 0 /protein complex that could be responsible for the hysteretic behavior. Both quinones inhibit the NADH: cytochrome b 5 reductase activity of human C b 5 R isoforms, but only CoQ 0 triggers hysteresis in the soluble C b 5 R isoform. Our data support the use of fully soluble components for studying hysteresis in this system, as they recapitulate key features of the behavior observed in biological membranes. • Cytochrome b 5 reductase activity of C b 5 R is inhibited by CoQ 0 and CoQ 10 • A lag phase emerges by soluble C b 5 R hysteretic modulation by CoQ 0 • The lag phase depends on soluble C b 5 R, C b 5 , and CoQ 0 concentration
Valério et al. (Sun,) studied this question.