Structural Plasticity and Ligand Promiscuity of CYP3A4 Revealed by Cryo-EM
- 1Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, CA 94158, USA
Abstract
Cytochrome P450 3A4 (CYP3A4) metabolizes roughly half of all marketed drugs, and its inhibition can cause clinically significant drug-drug interactions. The enzyme accommodates chemically diverse ligands, making binding modes and metabolic outcomes difficult to predict. Previous X-ray crystallography efforts have leveraged a truncated construct without the N-terminal segment that tethers CYP3A4 to the membrane. Here we show that the same construct assembles into a symmetric trimer that can be resolved by cryo-EM and determine structures of both unliganded and ligand-bound CYP3A4. Multiple ligands are resolved with density consistent with several mutually exclusive conformations. Protein remodeling to reshape the binding pocket is concentrated in the F/G loop, which is poorly resolved and unmodeled in many X-ray structures. These features likely underlie the poor predictive performance of co-folding methods on this target. The routine use of cryo-EM to resolve CYP3A4 ligand-bound complexes will provide the ground truth data needed to make predictive models of drug metabolism useful in practice.
Related
- Engineering catalytic promiscuity in serum albumin: synergistic roles of microenvironment, structural dynamics, and nano conjugation.
- Functional plasticity of AIF revealed by dimerization and CHCHD4 interaction states
- Structural diversity of human urinary N-glycans revealed by reversed-phase LC/MS/MS toward a resource for biomarker discovery
- Structural basis of phospholipid sensing nuclear receptor LRH-1 recognition by carboxylic acid bioisosteres reveals an alternative ligand interaction site
- Structural insights into MBOAT2 catalysis, product retention, and ligand exchange
BibTeX
@misc{orta2026structural,
title = {{Structural Plasticity and Ligand Promiscuity of CYP3A4 Revealed by Cryo-EM}},
author = {A. K. Orta and J.-H. Schäfer and G. J. Correy and J. O. Norman and J. Pampel and K. K. Huddleston and H. MacDermott-Opeskin and E. B. Miller and G. Reggiano and J. P. Rodrigues and G. C. Lander and W. P. Walters and J. S. Fraser},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.01.748687},
url = {https://www.biorxiv.org/content/10.64898/2026.09.01.748687v1},
note = {CC BY},
}
RIS
TY - UNPB
TI - Structural Plasticity and Ligand Promiscuity of CYP3A4 Revealed by Cryo-EM
AU - A. K. Orta
AU - J.-H. Schäfer
AU - G. J. Correy
AU - J. O. Norman
AU - J. Pampel
AU - K. K. Huddleston
AU - H. MacDermott-Opeskin
AU - E. B. Miller
AU - G. Reggiano
AU - J. P. Rodrigues
AU - G. C. Lander
AU - W. P. Walters
AU - J. S. Fraser
PY - 2026
DA - 2026/09/01
PB - bioRxiv
DO - 10.64898/2026.09.01.748687
UR - https://www.biorxiv.org/content/10.64898/2026.09.01.748687v1
AB - Cytochrome P450 3A4 (CYP3A4) metabolizes roughly half of all marketed drugs, and its inhibition can cause clinically significant drug-drug interactions. The enzyme accommodates chemically diverse ligands, making binding modes and metabolic outcomes difficult to predict. Previous X-ray crystallography efforts have leveraged a truncated construct without the N-terminal segment that tethers CYP3A4 to the membrane. Here we show that the same construct assembles into a symmetric trimer that can be resolved by cryo-EM and determine structures of both unliganded and ligand-bound CYP3A4. Multiple ligands are resolved with density consistent with several mutually exclusive conformations. Protein remodeling to reshape the binding pocket is concentrated in the F/G loop, which is poorly resolved and unmodeled in many X-ray structures. These features likely underlie the poor predictive performance of co-folding methods on this target. The routine use of cryo-EM to resolve CYP3A4 ligand-bound complexes will provide the ground truth data needed to make predictive models of drug metabolism useful in practice.
ER -