Direct interaction between the transmembrane helices stabilize cytochrome P450 2B4 and cytochrome b5 redox complex.
Biophysical chemistry(2023)
摘要
The catalytic activity of cytochrome P450 2B4 (CYP2B4) is moderated by its cognate redox partner cytochrome b5 (Cyt-b). The endoplasmic reticulum (ER) membrane and intermolecular transmembrane (TM) interaction between CYP2B4 and Cyt-b regulate the substrate catalysis and the reaction rate. This emphasizes the significance of elucidating the molecular basis of CYP2B4 and Cyt-b complexation in a membrane environment to better understand the enzymatic activity of CYP2B4. Our previous solid-state NMR studies revealed the membrane topology of the transmembrane domains of these proteins in the free and complex forms. Here, we show the cross-angle complex formation by the single-pass TM domains of CYP2B4 and Cyt-b, which is mainly driven by several salt-bridges (E2-R128, R21-D104 and K25-D104), using a multi-microsecond molecular dynamic simulation. Additionally, the leucine-zipper residues (L8, L12, L15, L18 and L19 from CYP2B4) and π-stacking between H23 and F20 residues of CYP2B4 and W110 of Cyt-b are identified to stabilize the TM-TM complex in the ER membrane. The simulated tilts of the helices in the free and in the complex are in excellent agreement with solid-state NMR results. The TM-TM packing influences a higher order structural stability when compared to the complex formed by the truncated soluble domains of these two proteins. MM/PBSA based binding free energy estimates nearly 100-fold higher binding affinity (ΔG = -2810.68 ± 696.44 kJ/mol) between the soluble domains of the full-length CYP2B4 and Cyt-b when embedded in lipid membrane as compared to the TM-domain-truncated soluble domains (ΔG = -27.406 ± 10.32 kJ/mol). The high-resolution full-length CYP2B4-Cyt-b complex structure and its dynamics in a native ER membrane environment reported here could aid in the development of approaches to effectively modulate the drug-metabolism activity of CYP2B4.
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关键词
transmembrane helices,p450
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