Bioactivation of chlorpyrifos by CYP2B6 variants

Abstract

1. Chlorpyrifos (CPF), an organophosphorus (OP) pesticide, is bioactivated by cytochrome P450s (CYPs) to the active metabolite chlorpyrifos oxon (CPF-O). Given that human CYP2B6 has the highest intrinsic clearance (CL_int) for CPF bioactivation, CYP2B6 polymorphisms may impact human susceptibility to CPF at real world environmental and occupational CPF exposure levels.

2. CYP2B6.4,.5,.7, and .18 were over-expressed in mammalian COS-1 cells to assess the impact of CYP2B6 variants on the K_m and V_max for bioactivation of CPF. Cell lysates were incubated with CPF (0–100 μM) and the production of CPF-O was measured via HPLC analysis. CYP2B6 content was determined by western blot.

3. CYP2B6.18 had neither detectable protein nor activity levels. The V_max value for each remaining variant was significantly higher than wild-type (CYP2B6.1, V_max 4.13 × 10⁴ pmol/min/nmol CYP2B6), with CYP2B6.4,.5, and .7 having V_max values of 4.52 × 10⁵, 1.82 × 10⁵, and 9.60 × 10⁴ pmol/min/nmol CYP2B6, respectively. The K_m values for these variants ranged from 0.39 to 1.09 μM and were not significantly different from wild-type. All active variants examined had significantly higher CL_int than CYP2B6.1.

4. Variants of CYP2B6 have altered capacity to bioactivate CPF and may affect individual susceptibility by altering the V_max for CPF-O formation.

Keywords::

• Biotransformation
• toxicokinetics
• pesticide metabolism
• cytochrome P450
• polymorphism