Anion-Dependent Stimulation of CYP3A4 Monooxygenase

Sevrioukova, Irina F.; Poulos, T.L.

doi:10.1021/acs.biochem.5b00510

Cited by 49 publications

(72 citation statements)

References 61 publications

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“…In the published structures of P450 1A1 and 1A2 complexed with α-naphthoflavone (PDB 2HI4, 4I8V) [37, 38], the main site of oxidation (for 5,6-epoxidation) [50] is furthest from the heme iron (12 Å). With (human) P450 3A4, some erythromycin-, bromoergocryptine-, and progesterone-bound structures have been published [18, 51–53] but only the bromoergocryptine complex (PDB 3XNU) is catalytically competent, i.e. the major sites of oxidation (8´ and 9´carbons of the proline ring) are 4.1 and 3.7 Å, respectively, away from the iron [52, 54].…”

Section: General Issues Of Active Site Flexibility and Multiple Occupmentioning

confidence: 99%

Recent Structural Insights into Cytochrome P450 Function

Guengerich

Waterman

Egli

2016

Trends in Pharmacological Sciences

290

178

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Cytochrome P450 (P450) enzymes are important in the metabolism of drugs, steroids, fat-soluble vitamins, carcinogens, pesticides, and many other types of chemicals. Their catalytic activities are important issues in areas such as drug-drug interactions and endocrine function. During the past 30 years, structures of P450s have been very helpful in understanding function, particularly the mammalian P450 structures available in the past 15 years. We review recent activity in this area, focusing on the past two years (2014–2015). Structural work with microbial P450s includes studies related to the biosynthesis of natural products and the use of parasitic and fungal P450 structures as targets for drug discovery. Studies on mammalian P450s include the utilization of information about ‘drug-metabolizing’ P450s to improve drug development and also to understand the molecular bases of endocrine dysfunction.

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Section: General Issues Of Active Site Flexibility and Multiple Occupmentioning

confidence: 99%

Recent Structural Insights into Cytochrome P450 Function

Guengerich

Waterman

Egli

2016

Trends in Pharmacological Sciences

290

178

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“…The intermediate TST binding site is reminiscent of that for progesterone (PRG) in crystal structures with CYP3A4; however there are important differences. (31,32) In Supporting Figure S1, an overlay of the intermediate structure with the CYP3A4-PRG crystal structure shows that PRG sits closer to the F'-helix and is packed against F213, F219 and F220. PRG is precluded from entering the active site by the interdigitation of the F108, F213, F215, F241 and F304 side chains.…”

Section: Accelerated Molecular Dynamicsmentioning

confidence: 99%

“…(Supporting Figure S2) Sevrioukova and Poulos recognized that the interface created by this interaction creates a hydrophobic patch to accommodate PRG. (32) This interaction apparently stabilizes the closed configuration of the Phe-cluster, precluding passage of this ligand into the active site. Assuming PRG enters the enzyme using the same mechanism as TST, it is unlikely that the PRG site represents a native transport intermediate, but rather a non-native one resulting from distortion of the native potential energy surface by the crystal lattice.…”

Section: Accelerated Molecular Dynamicsmentioning

confidence: 99%

Membrane-embedded substrate recognition by cytochrome P450 3A4

Hackett

2018

Journal of Biological Chemistry

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Cytochrome P450 3A4 (CYP3A4) is the dominant xenobiotic-metabolizing enzyme in the liver and intestine and is involved in the disposition of more than 50% of drugs. Owing to its ability to bind multiple substrates, its reaction kinetics are complex, and its association with the microsomal membrane confounds our understanding of how this enzyme recognizes and recruits diverse substrates. Testosterone (TST) hydroxylation is the prototypical CYP3A4 reaction, displaying positive homotropic cooperativity with three binding sites. Here, exploiting the capability of accelerated molecular dynamics (aMD) to sample events in the millisecond regime, I performed > 25-µs aMD simulations in the presence of three TST molecules. These simulations identified high-occupancy surfacebinding sites as well as a pathway for TST ingress into the CYP3A4 active site originating in the membrane. Adaptive biasing force analysis of the latter pathway revealed a metastable intermediate that could constitute a third binding site at high TST concentrations. Prompted by the observation that interactions between TST and the G'-helix mobilize the ligand into the active site, a free-energy analysis of TST distribution in the membrane was conducted and revealed that the depth of the G'-helix is optimal for extracting TST. In summary, these simulations confirm separate, but adjacent substrate-binding sites within the enzyme and the existence of an auxiliary TSTbinding site. The broader impact of these simulations is that they support a mechanism in which cytochromes P450 directly recruit membrane-solubilized substrates. ________________The cytochromes P450s metabolize nearly all drugs and environmental xenobiotics to which humans are exposed. Of the 57 P450s encoded by the human genome, cytochrome P450 3A4 (CYP3A4) is involved in the clearance of more than half of approved drugs and presents s i g n i f i c a n t c h a l l e n g e t o o p t i m i z i n g pharmacokinetics in drug development and avoiding adverse effects in the clinic.(1,2) CYP3A4 enzyme kinetics can demonstrate marked cooperativity.(3) Positive cooperativity is characterized by a continuous, positive increase in the rate of catalysis with increasing substrate concentration; whereas positive heterotropic cooperativity occurs when the rate of substrate turnover is enhanced by a second distinct effector molecule. Cooperativity in CYP catalysis was first recognized in microsome preparations(4-6) and later confirmed using purified CYP3A4 with several substrates including testosterone (TST), 17β-estradiol, amitriptyline, and aflatoxin B1. (7) A shift in the regioselectivity of midazolam (MDZ) oxidation in patients by concurrent fluconazole administration supports that heterotropic CYP3A4 interactions are also relevant in vivo.(8) Cooperativity in CYP3A4 catalysis has been explained by mechanisms invoking simultaneous occupation of the active site by multiple ligands or binding to nearby allosteric sites. (9)(10)(11)(12)(13)(14)(15) The membrane environment can have a profound effec...

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“…More recently, Sevrioukova and Poulos (2015) characterized a citrate binding site that was located near the F-G loop of CYP3A4. This is an area where anionic membrane phospholipids may interact with the P450.…”

Section: Anionic Phospholipidsmentioning

confidence: 99%

Cytochrome P450 Organization and Function Are Modulated by Endoplasmic Reticulum Phospholipid Heterogeneity

Brignac‐Huber¹,

Park²,

Reed³

et al. 2016

Drug Metabolism and Disposition

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Cytochrome P450s (P450s) comprise a superfamily of proteins that catalyze numerous monooxygenase reactions in animals, plants, and bacteria. In eukaryotic organisms, these proteins not only carry out reactions necessary for the metabolism of endogenous compounds, but they are also important in the oxidation of exogenous drugs and other foreign compounds. Eukaryotic P450 system proteins generally reside in membranes, primarily the endoplasmic reticulum or the mitochondrial membrane. These membranes provide a scaffold for the P450 system proteins that facilitate interactions with their redox partners as well as other P450s. This review focuses on the ability of specific lipid components to influence P450 activities, as well as the role of the membrane in P450 function. These studies have shown that P450s and NADPH-cytochrome P450 reductase appear to selectively associate with specific phospholipids and that these lipid-protein interactions influence P450 activities. Finally, because of the heterogeneous nature of the endoplasmic reticulum as well as other biologic membranes, the phospholipids are not arranged randomly but associate to generate lipid microdomains. Together, these characteristics can affect P450 function by 1) altering the conformation of the proteins, 2) influencing the P450 interactions with their redox partners, and 3) affecting the localization of the proteins into specific membrane microdomains.

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Anion-Dependent Stimulation of CYP3A4 Monooxygenase

Cited by 49 publications

References 61 publications

Recent Structural Insights into Cytochrome P450 Function

Recent Structural Insights into Cytochrome P450 Function

Membrane-embedded substrate recognition by cytochrome P450 3A4

Cytochrome P450 Organization and Function Are Modulated by Endoplasmic Reticulum Phospholipid Heterogeneity

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