Polymorphisms of CYP2C8 Alter First-Electron Transfer Kinetics and Increase Catalytic Uncoupling

Abstract

Cytochrome P450 2C8 (CYP2C8) epoxygenase is responsible for the metabolism of over 60 clinically relevant drugs, notably the anticancer drug Taxol (paclitaxel, PAC). Specifically, there are naturally occurring polymorphisms, CYP2C8*2 and CYP2C8*3, that display altered PAC hydroxylation rates despite these mutations not being located in the active site. Herein, we demonstrate that these polymorphisms result in a greater uncoupling of PAC metabolism by increasing the amount of hydrogen peroxide formed per PAC turnover. Anaerobic stopped-flow measurements determined that these polymorphisms have altered first electron transfer kinetics, compared to CYP2C8*1 (wildtype), that suggest electron transfer from cytochrome P450 reductase (CPR) is disfavored. Therefore, these data demonstrate that these polymorphisms affect the catalytic cycle of CYP2C8 and suggest that redox interactions with CPR are disrupted.

A disease‐associated missense mutation in CYP4F3 affects the metabolism of leukotriene B4 via disruption of electron transfer

“…We propose that L375 is important for stabilizing a nearby α‐helix which carries the crucial residues for the electron shuffling process, and that the L375V mutation destabilizes the helix‐mediating electron shuffling, thereby disrupting the catalytic function of CYP4F3. Point mutations affecting electron transfer in CYP enzymes have been reported before, such as polymorphisms in CYP2C8 affecting its hydroxylase function 40 . The same goes for variations in electron transfer partners that disrupt CYP activity 41 …”

“…Point mutations affecting electron transfer in CYP enzymes have been reported before, such as polymorphisms in CYP2C8 affecting its hydroxylase function. 40 The same goes for variations in electron transfer partners that disrupt CYP activity. 41 …”

Role of cytochrome P450 2C8 genetic polymorphism and epoxygenase uncoupling in periodontal remodelling affecting orthodontic treatment

“…Being capable of producing reactive oxygen species, cytochrome enzymes may contribute to the cellular oxidation–reduction balance through a process in their reaction cycle called uncoupling 15 . Uncoupling describes the formation of ROS by decomposition of reaction intermediates instead of completing substrate oxidation 5 . It is suggested that uncoupling can have damaging effects by contributing to the cellular ROS levels 16 .…”

“…15 Uncoupling describes the formation of ROS by decomposition of reaction intermediates instead of completing substrate oxidation. 5 It is suggested that uncoupling can have damaging effects by contributing to the cellular ROS levels. 16 We measured H 2 O 2 generation by isolated CYP2C8 enzymes and detected an increased formation in the CYP2C8*3 variant enzyme enhanced by zoledronic acid and also by arachidonic acid (Figure 4C,F).…”

“…It has been shown that different allelic variants of the CYP2C8 gene have differing effects on substrate metabolism. While substrate metabolism was only slightly decreased in CYP2C8*2 and *3 allelic variant enzymes, it has been shown that these variants are prone to the unfavourable side reaction of uncoupling, resulting in the direct release of reactive oxygen species, without completing substrate oxidation 5,6 . Specifically, the generated iron–oxygen decays before substrate oxidation has been completed; superoxide or hydrogen peroxide can be formed as toxic side products.…”