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Conformational flexibility of mammalian cytochrome P450 2B4 in binding imidazole inhibitors of different ring chemistry and side chains
Author(s) -
Muralidhara Bilikallahalli,
Negi Surendra,
Chin Christopher C,
Braun Werner,
Halpert James R
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.4.a264
Subject(s) - isothermal titration calorimetry , chemistry , imidazole , enthalpy , stereochemistry , titration , crystallography , thermodynamics , physics
Recent x‐ray structures of cytochrome P450 2B4 (CYP2B4) reveal an open form that undergoes a large‐scale structural transition to a closed form upon 4‐(4‐chlorophenyl)imidazole (4‐CPI) binding. Here we report for the first time using isothermal titration calorimetry (ITC) and spectroscopic titrations the conformational flexibility of CYP2B4 in solution upon binding six different inhibitors:4‐CPI, 1‐benzylimidazole, 4‐phenylimidazole, 1‐(4‐chlorophenyl)imidazole, 1‐phenylimidazole and 1‐(2‐(benzyloxy)ethyl)imidazole (BEI). All the inhibitors induce type‐II spectral changes. Calorimetric titrations using monomeric enzyme yielded 1:1 binding stoichiometry with the associated K D values ranging from 0.2 μM to 4.8 μM and changes in enthalpy from −6.5 to −8.8 kcal mol −1 . The largest difference in entropy (+5.9 vs. −4.1 cal mol −1 K −1 ) and heat capacity change (−604 vs. −331 cal mol −1 K −1 ) was observed between 4‐CPI and BEI. Interaction of all six inhibitors was enthalpically driven and hydrophobic, and only 4‐CPI induces large conformational change in the enzyme. Accessibility to acrylamide of the only tryptophan (Trp 121 ) and molecular modeling studies complement these findings. The thermodynamic signature obtained from ITC can be an excellent resource for minimizing deleterious P450 interactions in rational drug design and for picking compounds for structural analysis of P450 enzymes. NCAA Grant MCB050038N (BKM), NIH Grant ES03619, and Center grant ES06676 (J.R.H.).