Cooperative effects for CYP2E1 differ between styrene and its metabolites

Hartman, Jessica H.; Boysen, Gunnar; Miller, Grover P.

doi:10.3109/00498254.2012.760764

Cited by 11 publications

(28 citation statements)

References 29 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…For 4-nitrophenol, metabolic rates of turnover increase and then decrease as a function of substrate concentration indicating substrate inhibition [3, 6]. Alternatively, many CYP2E1 substrates, including phenacetin, m -xylene [5], styrene [7, 8], and 7-ethoxycoumarin, demonstrate a poor efficiency in turnover at low substrate concentrations that rapidly improves at higher concentrations through a positive cooperative mechanism. Recent studies have further shown that aniline metabolism by CYP2E1 metabolism involves negative cooperativity in which higher substrate concentrations inhibit the ability for the enzyme to reach a maximal rate [9].…”

Section: Introductionmentioning

confidence: 99%

Structural basis for cooperative binding of azoles to CYP2E1 as interpreted through guided molecular dynamics simulations

Levy

Hartman

Perry

et al. 2015

Journal of Molecular Graphics and Modelling

Self Cite

View full text Add to dashboard Cite

CYP2E1 metabolizes a wide array of small, hydrophobic molecules, resulting in their detoxification or activation into carcinogens through Michaelis-Menten as well as cooperative mechanisms. Nevertheless, the molecular determinants for CYP2E1 specificity and metabolic efficiency toward these compounds are still unknown. Herein, we employed computational docking studies coupled to Molecular Dynamics simulations to provide a critical perspective for understanding a structural basis for cooperativity observed for an array of azoles from our previous binding and catalytic studies (Hartman, JH et al (2014) Biochem Pharmacol 87, 523-33). The resulting 28 CYP2E1 complexes in this study revealed a common passageway for azoles that included a hydrophobic steric barrier causing a pause in movement toward the active site. The entrance to the active site acted like a second sieve to restrict access to the inner chamber. Collectively, these interactions impacted the final orientation of azoles reaching the active site and hence could explain differences in their biochemical properties observed in our previous studies, such as the consequences of methylation at position 5 of the azole ring. The association of a second azole demonstrated significant differences in interactions stabilizing the bound complex than observed for the first binding event. Intermolecular interactions occurred between the two azoles as well as CYP2E1 residue side chains and backbone and involved both hydrophobic contacts and hydrogen bonds. The relative importance of these interactions depended on the structure of the respective azoles indicating the absence of specific defining criteria for binding unlike the well-characterized dominant role of hydrophobicity in active site binding. Consequently, the structure activity relationships described here and elsewhere are necessary to more accurately identify factors impacting the observation and significance of cooperativity in CYP2E1 binding and catalysis toward drugs, dietary compounds, and pollutants.

show abstract

Section: Introductionmentioning

confidence: 99%

Structural basis for cooperative binding of azoles to CYP2E1 as interpreted through guided molecular dynamics simulations

Levy

Hartman

Perry

et al. 2015

Journal of Molecular Graphics and Modelling

Self Cite

View full text Add to dashboard Cite

show abstract

“…These studies were followed up by work from other groups establishing a structural basis for CYP2E1 selectivity toward molecules [12, 13]. Moreover, we recently showed through catalytic studies with styrene and its metabolites that the structure of the molecules influenced their potency and mechanism of inhibition toward 4-nitrophenol metabolism [14]. Styrene and its primary oxidized metabolites, styrene oxide and 4-vinylphenol, differed little in hydrophobicity and shared similar affinities for both CYP2E1 binding sites.…”

Section: Introductionmentioning

confidence: 99%

Structure of pyrazole derivatives impact their affinity, stoichiometry, and cooperative interactions for CYP2E1 complexes

Hartman¹,

Bradley²,

Laddusaw³

et al. 2013

Archives of Biochemistry and Biophysics

Self Cite

View full text Add to dashboard Cite

CYP2E1 plays a critical role in detoxication and carcinogenic activation of drugs, pollutants, and dietary compounds; however, these metabolic processes can involve poorly characterized cooperative interactions that compromise the ability to understand and predict CYP2E1 metabolism. Herein, we employed an array of ten azoles with an emphasis on pyrazoles to establish the selectivity of catalytic and cooperative CYP2E1 sites through binding and catalytic studies. Spectral binding studies for monocyclic azoles suggested two binding events, while bicyclic azoles suggested one. Pyrazole had moderate affinity toward the CYP2E1 catalytic site that improved when a methyl group was introduced at either position 3 or 4. The presence of methyl groups simultaneously at positions 3 and 5 blocked binding, and a phenyl group at position 3 did not improve binding affinity. In contrast, pyrazole fusion to a benzene or cyclohexane ring greatly increased affinity. The consequences of these binding events on CYP2E1 catalysis were studied through inhibition studies with 4-nitrophenol, a substrate known to bind both sites. Most pyrazoles shared a common mixed cooperative inhibition mechanism in which pyrazole binding rescued CYP2E1 from substrate inhibition. Overall, inhibitor affinities toward the CYP2E1 catalytic site were similar to those reported in binding studies, and the same trend was observed for binding at the cooperative site. Taken together, these studies identified key structural determinants in the affinity and stoichiometry of azole interactions with CYP2E1 and consequences on catalysis that further advance an understanding of the relationship between structure and function for this enzyme.

show abstract

“…Acetaminophen acted as a mixed-type inhibitor on the styrene reaction, which was similar to inhibition mechanisms reported for many other five- and six- member aromatic and heterocyclic compounds. 4, 7, 8, 12 In particular, there were many similarities between the effects of acetaminophen on styrene epoxidation to that by 4-methylpyrazole. 4 First, acetaminophen bound to the binary (enzyme-styrene) complex, yet CYP2E1 remained active toward styrene, albeit with a slightly lower rate (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…8, red circles), 4, 7 and thus it was possible to fully interrogate the impact of acetaminophen on styrene conversion to a reactive epoxide metabolite. Acetaminophen acted as a mixed-type inhibitor on the styrene reaction, which was similar to inhibition mechanisms reported for many other five- and six- member aromatic and heterocyclic compounds.…”

Section: Discussionmentioning

confidence: 99%

Cooperativity in CYP2E1 metabolism of acetaminophen and styrene mixtures

Hartman

Letzig

Roberts

et al. 2015

Biochemical Pharmacology

Self Cite

View full text Add to dashboard Cite

Risk assessment for exposure to mixtures of drugs and pollutants relies heavily on in vitro characterization of their bioactivation and/or metabolism individually and extrapolation to mixtures assuming no interaction. Herein, we demonstrated that in vitro CYP2E1 metabolic activation of acetaminophen and styrene mixtures could not be explained through the Michaelis-Menten mechanism or any models relying on that premise. As a baseline for mixture studies with styrene, steady-state analysis of acetaminophen oxidation revealed a biphasic kinetic profile that was best described by negative cooperativity (Hill coefficient = 0.72). The best-fit mechanism for this relationship involved two binding sites with differing affinities (Ks = 830 µM and Kss = 32 mM). Introduction of styrene inhibited that reaction less than predicted by simple competition and thus provided evidence for a cooperative mechanism within the mixture. Likewise, acetaminophen acted through a mixed-type inhibition mechanism to impact styrene epoxidation. In this case, acetaminophen competed with styrene for CYP2E1 (Ki = 830 µM and Ksi = 180 µM for catalytic and effector sites, respectively) and resulted in cooperative impacts on binding and catalysis. Based on modeling of in vivo clearance, cooperative interactions between acetaminophen and styrene resulted in profoundly increased styrene activation at low styrene exposure levels and therapeutic acetaminophen levels. Current Michaelis-Menten based toxicological models for mixtures such as styrene and acetaminophen would fail to detect this concentration-dependent relationship. Hence, future studies must assess the role of alternate CYP2E1 mechanisms in bioactivation of compounds to improve the accuracy of interpretations and predictions of toxicity.

show abstract

Cooperative effects for CYP2E1 differ between styrene and its metabolites

Cited by 11 publications

References 29 publications

Structural basis for cooperative binding of azoles to CYP2E1 as interpreted through guided molecular dynamics simulations

Structural basis for cooperative binding of azoles to CYP2E1 as interpreted through guided molecular dynamics simulations

Structure of pyrazole derivatives impact their affinity, stoichiometry, and cooperative interactions for CYP2E1 complexes

Cooperativity in CYP2E1 metabolism of acetaminophen and styrene mixtures

Contact Info

Product

Resources

About