Evaluation of Atypical Cytochrome P450 Kinetics with Two-Substrate Models:  Evidence That Multiple Substrates Can Simultaneously Bind to Cytochrome P450 Active Sites†

Substrate specificity and selectivity of a biocatalyst are determined by the protein sequence and structure of its active site. Finding versatile biocatalysts acting against multiple substrates while at the same time being chiral selective is of interest for the pharmaceutical and chemical industry. However, the relationships between these two properties in natural microbial enzymes remain underexplored. Here, we performed an experimental analysis of substrate promiscuity and chiral selectivity in a set of 145 purified esterases from phylogenetically and environmentally diverse microorganisms, which were assayed against 96 diverse esters, 20 of which were enantiomers. Our results revealed a negative correlation between substrate promiscuity and chiral selectivity in the evaluated enzymes. Esterases displaying prominent substrate promiscuity and large catalytic environments are characterized by low chiral selectivity, a feature that has limited commercial value. Although a low level of substrate promiscuity does not guarantee high chiral selectivity, the probability that esterases with smaller active sites possess chiral selectivity factors of interest for industry (>25) is significantly higher than for promiscuous enzymes. Together, the present study unambiguously demonstrates that promiscuous and selective esterases appear to be rare in nature and that substrate promiscuity can be used as an indicator of the chiral selectivity level of esterases, and vice versa.

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A Computational Approach towards the Development of Newer Anticancer Agents

E-Journal of Chemistry ◽

10.1155/2010/582710 ◽

2010 ◽

Vol 7 (1) ◽

pp. 21-24 ◽

Cited By ~ 1

Author(s):

Javed I. Sheikh ◽

H. D. Juneja ◽

V. N. Ingle

Keyword(s):

Cytochrome P450 ◽

Molecular Docking ◽

Anticancer Agents ◽

In Silico ◽

Active Sites ◽

Computational Approach ◽

The Novel ◽

Cytochrome P450 Reductase ◽

P450 Reductase

In silicomolecular docking analyses of the novel 1-(2', 4'-dihydroxy-5'-chlorophenyl)-3-aryl-propane-1, 3-diones were performed in the active sites of enzyme cytochrome P450 reductase to obtain new anticancer agents computationally. 1-(2', 4'-dihydroxy-5'-chlorophenyl)-3-(4"-methoxy-phenyl)-propane-1, 3-dione(3f)was found to have maximum affinity for the active sites of enzyme.

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Stereospecific Activation of the Procarcinogen Benzo[a]pyrene: A Probe for the Active Sites of the Cytochrome P450 Superfamily

Biochemistry ◽

10.1021/bi00021a007 ◽

1995 ◽

Vol 34 (21) ◽

pp. 6956-6961 ◽

Cited By ~ 15

Author(s):

Jeffrey P. Jones ◽

Magang Shou ◽

Kenneth R. Korzekwa

Keyword(s):

Cytochrome P450 ◽

Active Sites

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MEMS directed evolution of two cytochrome P450 enzymes revealing distinct active-sites for convergent functions

10.1101/2020.12.25.424376 ◽

2020 ◽

Author(s):

Li Ma ◽

Fengwei Li ◽

Xingwang Zhang ◽

Hui Chen ◽

Qian Huang ◽

...

Keyword(s):

Cytochrome P450 ◽

Directed Evolution ◽

Active Sites ◽

Enzyme Engineering ◽

Proof Of Concept ◽

Cytochrome P450 Enzymes ◽

Natural Evolution ◽

One Pot ◽

P450 Bm3 ◽

Protein Enzyme

AbstractDirected evolution (DE) inspired by natural evolution (NE) has been achieving tremendous successes in protein/enzyme engineering. However, the conventional ‘one-protein-for-one-task’ DE cannot match the ‘multi-proteins-for-multi-tasks’ NE in terms of screening throughput and efficiency, thus often failing to meet the fast-growing demands for biocatalysts with desired properties. In this study, we design a novel ‘multi-enzyme-for-multi-substrate’ (MEMS) DE model and establish the proof-of-concept by running a NE-mimicking and higher-throughput screening on the basis of ‘two-P450s-against-seven-substrates’ (2P×7S) in one pot. With the significantly improved throughput and hit-rate, we witness a series of convergent evolution events of the two archetypal cytochrome P450 enzymes (P450 BM3 and P450cam) in laboratory. Further structural analysis of the two functionally convergent P450 variants provide important insights into how distinct active-sites can reach a common catalytic goal.

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Bioactivity of curcumin on the cytochrome P450 enzymes of the steroidogenic pathway

10.1101/669440 ◽

2019 ◽

Cited By ~ 1

Author(s):

Patricia Rodríguez Castaño ◽

Shaheena Parween ◽

Amit V Pandey

Keyword(s):

Cytochrome P450 ◽

Active Sites ◽

Curcuma Longa ◽

Docking Studies ◽

Steroid Metabolism ◽

Dependent Manner ◽

Cytochrome P450 Enzymes ◽

Lead Compounds ◽

Anti Cancer ◽

Mild Reduction

AbstractTurmeric, a popular ingredient in the cuisine of many Asian countries, comes from the roots of theCurcuma longaand is known for its use in Chinese and Ayurvedic medicine. Turmeric is rich in curcuminoids, including curcumin, demethoxycurcumin, and bisdemethoxycurcumin. Curcuminoids have potent wound healing, anti-inflammatory, and anti-carcinogenic activities. While curcuminoids have been studied for many years, not much is known about their effects on steroid metabolism. Since many anti-cancer drugs target enzymes from the steroidogenic pathway, we tested the effect of curcuminoids on cytochrome P450 CYP17A1, CYP21A2, and CYP19A1 enzyme activities. When using 10 µg/ml of curcuminoids, both the 17α-hydroxylase as well as 17,20 lyase activities of CYP17A1 were reduced significantly. On the other hand, only a mild reduction in CYP21A2 activity was observed. Furthermore, CYP19A1 activity was also reduced up to ~20% of control when using 1-100 µg/ml of curcuminoids in a dose-dependent manner. Molecular docking studies confirmed that curcumin could dock into the active sites of CYP17A1, CYP19A1 as well as CYP21A2. In CYP17A1 and CYP19A1, curcumin docked within 2.5 Å of central heme while in CYP21A2 the distance from heme was 3.4 Å, which is still in the same range or lower than distances of bound steroid substrates. These studies suggest that curcuminoids may cause inhibition of steroid metabolism, especially at higher dosages. Also, the recent popularity of turmeric powder as a dilatory supplement needs further evaluation for the effect of curcuminoids on steroid metabolism. Molecular structure of curcuminoids could be modified to generate better lead compounds with inhibitory effects on CYP17A1 and CYP19A1 for potential drugs against prostate cancer and breast cancer.

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Cytochrome P450 in GtoPdb v.2021.2

IUPHAR/BPS Guide to Pharmacology CITE ◽

10.2218/gtopdb/f242/2021.2 ◽

2021 ◽

Vol 2021 (2) ◽

Author(s):

Kathryn Burns ◽

Nuala Ann Helsby

Keyword(s):

Cytochrome P450 ◽

Drug Disposition ◽

Cytochrome P450 Enzyme ◽

Multiple Substrates ◽

Cyp Enzymes ◽

Physiological Processes ◽

Enzyme Superfamily ◽

P450 Enzyme ◽

The Individual ◽

Fat Soluble Vitamins

The cytochrome P450 enzyme superfamily (CYP), E.C. 1.14.-.-, are haem-containing monooxygenases with a vast range of both endogenous and exogenous substrates. These include sterols, fatty acids, eicosanoids, fat-soluble vitamins, hormones, pesticides and carcinogens as well as drugs. Listed below are the human enzymes, their relationship with rodent CYP enzyme activities is obscure in that the species orthologue may not metabolise the same substrates. Some of the CYP enzymes located in the liver are particularly important for drug metabolism, both hepatic and extrahepatic CYP enzymes also contribute to patho/physiological processes. Genetic variation of CYP isoforms is widespread and likely underlies a proportion of individual variation in drug disposition. The superfamily has the root symbol CYP, followed by a number to indicate the family, a capital letter for the subfamily with a numeral for the individual enzyme. Some CYP are able to metabolise multiple substrates, others are oligo- or mono- specific.

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Malleability and Versatility of Cytochrome P450 Active Sites Studied by Molecular Simulations

Current Drug Metabolism ◽

10.2174/138920012798918453 ◽

2012 ◽

Vol 13 (2) ◽

pp. 190-196 ◽

Cited By ~ 14

Author(s):

Chris Oostenbrink ◽

Anita de Ruiter ◽

Jozef Hritz ◽

Nico Vermeulen

Keyword(s):

Cytochrome P450 ◽

Active Sites ◽

Molecular Simulations

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Cytochrome P450 2D6.1 and cytochrome P450 2D6.10 differ in catalytic activity for multiple substrates

Pharmacogenetics ◽

10.1097/00008571-200108000-00003 ◽

2001 ◽

Vol 11 (6) ◽

pp. 477-487 ◽

Cited By ~ 68

Author(s):

Yamini Ramamoorthy ◽

Rachel F. Tyndale ◽

Edward M. Sellers

Keyword(s):

Cytochrome P450 ◽

Catalytic Activity ◽

Multiple Substrates

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Cytochrome P450 and P-gp Mediated Herb-Drug Interactions and Molecular Docking Studies of Garcinol

Membranes ◽

10.3390/membranes11120992 ◽

2021 ◽

Vol 11 (12) ◽

pp. 992

Author(s):

Lavanya Bolla ◽

Pratima Srivastava ◽

Velayutham Ravichandiran ◽

Satheesh Kumar Nanjappan

Keyword(s):

Cytochrome P450 ◽

Molecular Docking ◽

Drug Interactions ◽

Active Sites ◽

Docking Studies ◽

Rat Liver Microsomes ◽

Garcinia Cambogia ◽

Molecular Docking Studies ◽

Garcinia Indica

Garcinol is an active constituent of Garcinia indica and Garcinia cambogia. Recent studies have proven that garcinol has anti-inflammatory, anti-cancer, and anti-oxidant activities. The objective of this study was to evaluate the inhibitory effects of garcinol on the activities of the drug metabolizing cytochrome P450 (CYP) isozymes to predict potential herb-drug interactions with co-administered drugs. Garcinol was incubated with a mixture of rat liver microsomes and eight CYP probe substrate cocktail under optimized incubation conditions and the samples were analyzed using a validated method on LC-MS/MS. Garcinol showed strong inhibition with IC50 values of CYP1A2 (7.6 µM), CYP2C9 (8.0 µM), CYP2B6 (2.1 µM), CYP2D6 (9.5 µM), and CYP3A4 (5.1 µM), respectively, and moderate inhibition towards CYP2C19 (16.4 µM) and CYP2E1 (19.0 µM). Molecular docking studies were performed on garcinol against the active sites of CYP2B6 and CYP3A4 proteins. These results further confirmed that the inhibitory activity of garcinol occurred by occupying the active sites of these human CYPs and by making favorable interactions with its key residues. In-vivo CYP inhibition studies were carried out in Sprague-Dawley rats. These results suggest garcinol may cause herb-drug interactions, mediated by inhibition of CYPs involved in drug metabolism in-vivo by altering the pharmacokinetic parameters like AUC and Cmax in a clinically significant manner. Garcinol was found to upregulate the expression and activity of P-gp in western blotting study and P-gp inhibition study in-vivo. These findings give a clear understanding to predict potential herb-drug/drug-drug interactions of garcinol for safe clinical use in future.

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