Structural Diversity of Eukaryotic Membrane Cytochrome P450s

Johnson, Eric F.; Stout, C.D.

doi:10.1074/jbc.r113.452805

Cited by 94 publications

(83 citation statements)

References 101 publications

(89 reference statements)

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“…As shown in Fig. 4C, the amino acid residues lining the active-site cavity as well as adjacent residues of rabbit P450 4B1 are highly conserved for other characterized mammalian P450 4B1's, in sharp contrast to the high degree of active site divergence evident for family 2 paralogues that contribute to a broad capacity for xenobiotic metabolism (47). Moreover, the residues that position the substrate near the heme iron (labeled with bold text in Fig.…”

Section: Resultsmentioning

confidence: 92%

The Crystal Structure of Cytochrome P450 4B1 (CYP4B1) Monooxygenase Complexed with Octane Discloses Several Structural Adaptations for ω-Hydroxylation

Hsu

Baer

Rettie

et al. 2017

Journal of Biological Chemistry

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Edited by Ruma BanerjeeP450 family 4 fatty acid -hydroxylases preferentially oxygenate primary C-H bonds over adjacent, energetically favored secondary C-H bonds, but the mechanism explaining this intriguing preference is unclear. To this end, the structure of rabbit P450 4B1 complexed with its substrate octane was determined by X-ray crystallography to define features of the active site that contribute to a preference for -hydroxylation. The structure indicated that octane is bound in a narrow active-site cavity that limits access of the secondary C-H bond to the reactive intermediate. A highly conserved sequence motif on helix I contributes to positioning the terminal carbon of octane for -hydroxylation. Glu-310 of this motif auto-catalytically forms an ester bond with the heme 5-methyl, and the immobilized Glu-310 contributes to substrate positioning. The preference for -hydroxylation was decreased in an E310A mutant having a shorter side chain, but the overall rates of metabolism were retained. E310D and E310Q substitutions having longer side chains exhibit lower overall rates, likely due to higher conformational entropy for these residues, but they retained high preferences for octane -hydroxylation. Sequence comparisons indicated that active-site residues constraining octane for -hydroxylation are conserved in family 4 P450s. Moreover, the heme 7-propionate is positioned in the active site and provides additional restraints on substrate binding. In conclusion, P450 4B1 exhibits structural adaptations for -hydroxylation that include changes in the conformation of the heme and changes in a highly conserved helix I motif that is associated with selective oxygenation of unactivated primary C-H bonds.Cytochrome P450 family 4 fatty acid -hydroxylases are heme-containing monooxygenases that provide pathways for the reduction of potentially toxic non-esterified fatty acids and for the elimination of excess nutritive and non-nutritive lipids by catalyzing the addition of oxygen to a C-H bond of the terminal -carbon of fatty acids (1, 2). The resulting -alcohols are generally oxidized further to produce dicarboxylic acids, but they also may be transformed to glucuronides or esterified to glycerolipids. Urinary dicarboxylic fatty acids are elevated in humans by fasting or in uncontrolled diabetes, which are conditions where adipocyte lipolysis increases the availability of non-esterified fatty acids to fuel metabolism (3). It is estimated that roughly 15% of fatty acids undergo -hydroxylation during peak periods of fatty acid catabolism (4), whereas this fraction is much smaller under conditions where concentrations of nonesterified fatty acids are low (5). Collectively, these enzymes provide pathways for the metabolic clearance of branched chain fatty acids, eicosanoids, and xenobiotic substrates such as dietary phytanic acid, drugs, toxins, and vitamins E and K (1, 2, 6).Similar to other P450 2 gene families encoding multipurpose enzymes for the elimination of xenobiotic compounds, family 4 exhibits genetic diversi...

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Section: Resultsmentioning

confidence: 92%

The Crystal Structure of Cytochrome P450 4B1 (CYP4B1) Monooxygenase Complexed with Octane Discloses Several Structural Adaptations for ω-Hydroxylation

Hsu

Baer

Rettie

et al. 2017

Journal of Biological Chemistry

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“…The majority of eukaryotic CYPs are cotranslationally inserted into the ER membrane and remain anchored by an N-terminal hydrophobic helix (Chapple, 1998). The N-terminal leader includes the transmembrane domain (Johnson and Stout, 2013). However, CYPs are not restricted exclusively to this subcellular location.…”

Section: Discussionmentioning

confidence: 99%

Biphenyl 4-Hydroxylases Involved in Aucuparin Biosynthesis in Rowan and Apple Are Cytochrome P450 736A Proteins

Sircar¹,

Gaid

Chizzali

et al. 2015

Plant Physiol.

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Upon pathogen attack, fruit trees such as apple (Malus spp.) and pear (Pyrus spp.) accumulate biphenyl and dibenzofuran phytoalexins, with aucuparin as a major biphenyl compound. 4-Hydroxylation of the biphenyl scaffold, formed by biphenyl synthase (BIS), is catalyzed by a cytochrome P450 (CYP). The biphenyl 4-hydroxylase (B4H) coding sequence of rowan (Sorbus aucuparia) was isolated and functionally expressed in yeast (Saccharomyces cerevisiae). SaB4H was named CYP736A107. No catalytic function of CYP736 was known previously. SaB4H exhibited absolute specificity for 3-hydroxy-5-methoxybiphenyl. In rowan cell cultures treated with elicitor from the scab fungus, transient increases in the SaB4H, SaBIS, and phenylalanine ammonia lyase transcript levels preceded phytoalexin accumulation. Transient expression of a carboxyl-terminal reporter gene construct directed SaB4H to the endoplasmic reticulum. A construct lacking the amino-terminal leader and transmembrane domain caused cytoplasmic localization. Functional B4H coding sequences were also isolated from two apple (Malus 3 domestica) cultivars. The MdB4Hs were named CYP736A163. When stems of cv Golden Delicious were infected with the fire blight bacterium, highest MdB4H transcript levels were observed in the transition zone. In a phylogenetic tree, the three B4Hs were closest to coniferaldehyde 5-hydroxylases involved in lignin biosynthesis, suggesting a common ancestor. Coniferaldehyde and related compounds were not converted by SaB4H.

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“…The best model was aligned with the available CYP2C8 structure complexed with felodipine (Protein Data Bank code 2NNJ) using the PyMOL Molecular Graphics System (Version 1.5.0.4, Schrödinger). The putative heme-binding residues of WTCY2C8 and Var_3 were predicted based on the known structures of CYP2C5 and CYP2C8 (33,37,38).…”

Section: Isolation Of Mitochondria and Microsomes From Frozenmentioning

confidence: 99%

“…Second, a nearly full-length form (comparable in size with the microsomal CYP2C8; designated as Var_1) and a truncated form (44 kDa; designated as Var_3) were detected in mitochondria. Notably, despite missing the critical Arg-97 residue involved in binding to the heme group in the full-length enzyme (33), the N-terminal truncated form is capable of binding to heme and catalyzed the metabolism of some but not all CYP2C8 substrates. In this study, we show that three major splice variants of CYP2C8 expressed in human liver exhibit remarkable differences with respect to relative levels of microsomal (ER) and mitochondrial targeting, metabolism of paclitaxel and arachidonic acid, and induction of cellular oxidative stress.…”

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confidence: 99%

Targeting of Splice Variants of Human Cytochrome P450 2C8 (CYP2C8) to Mitochondria and Their Role in Arachidonic Acid Metabolism and Respiratory Dysfunction

Bajpai

Srinivasan

Ghosh

et al. 2014

Journal of Biological Chemistry

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Structural Diversity of Eukaryotic Membrane Cytochrome P450s

Cited by 94 publications

References 101 publications

The Crystal Structure of Cytochrome P450 4B1 (CYP4B1) Monooxygenase Complexed with Octane Discloses Several Structural Adaptations for ω-Hydroxylation

The Crystal Structure of Cytochrome P450 4B1 (CYP4B1) Monooxygenase Complexed with Octane Discloses Several Structural Adaptations for ω-Hydroxylation

Biphenyl 4-Hydroxylases Involved in Aucuparin Biosynthesis in Rowan and Apple Are Cytochrome P450 736A Proteins

Targeting of Splice Variants of Human Cytochrome P450 2C8 (CYP2C8) to Mitochondria and Their Role in Arachidonic Acid Metabolism and Respiratory Dysfunction

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