Residue 208 of UGT1A7 Determines the Functional Consequences of CYP3A4-UGT1A7 Association in Living Cells

Cytochrome P450 3A4 (CYP3A4) and UDP-glucuronosyltransferase 1A7 (UGT1A7) are known to interact functionally on the endoplasmic reticulum membrane. In a previous study, CYP3A4 enhanced the activities of UGT1A7*1 and UGT1A7*2 but suppressed that of UGT1A7*3, suggesting a role for residue 208 in CYP3A4-dependent regulation. To test this hypothesis, we investigated UGT1A7*4, a naturally occurring variant carrying the W208R substitution. Enzyme kinetic analyses demonstrated that UGT1A7*4 retained basal catalytic activity comparable with that of wild-type UGT1A7*1. However, co-expression of CYP3A4 suppressed UGT1A7*4 activity, whereas it enhanced UGT1A7*1 activity. This opposite regulatory response was consistently observed with 4-methylumbelliferone (4-MU), SN-38, and 4-hydroxybiphenyl (4-OHBP). Pull-down assays showed that UGT1A7*1, UGT1A7*4, and a W208A mutant all retained detectable association with CYP3A4. FRET analysis further demonstrated direct interaction between CYP3A4 and both UGT1A7 variants in living cells. These findings indicate that residue 208 influences the functional consequences of CYP3A4-mediated regulation without abolishing physical association between CYP3A4 and UGT1A7. We propose that residue 208 may function as a determinant, and potentially a molecular switch, governing the outcome of CYP3A4-dependent modulation of UGT1A7 activity.