scholarly journals Morphological adrenarche in rhesus macaques: development of the zona reticularis is concurrent with fetal zone regression in the early neonatal period

2008 ◽  
Vol 199 (3) ◽  
pp. 367-378 ◽  
Author(s):  
Ann D Nguyen ◽  
Samantha M Mapes ◽  
C Jo Corbin ◽  
Alan J Conley
Keyword(s):  
Cytochrome P450 ◽  
Rhesus Macaques ◽  
Expression Profiles ◽  
Cytochrome B5 ◽  
Zona Reticularis ◽  
Redox Partner ◽  
Cytochrome P450 Oxidoreductase ◽  
Dense Band ◽  
Cytochrome P450 Family ◽  
Steroid Dehydrogenase

Human adrenarche is associated with the establishment of a functional zona reticularis (ZR) and increasing secretion of dehydroepiandrosterone (DHEA) in sulfated form (DS). Like most non-human primates, rhesus macaques are not believed to undergo adrenarche, though they clearly establish a functional ZR after birth. However, the origins of the rhesus ZR are not well defined. Therefore, we investigated the zonal development, steroidogenic enzyme expression and morphology of rhesus adrenals from 1 day to 14 months of age. Immunohistochemistry was conducted to determine expression profiles of the steroidogenic enzymes 17α-hydroxylase/17,20-lyase cytochrome P450, family 17, subfamily A, polypeptide 1 (CYP17A1), cytochrome P450, family 21, subfamily A, polypeptide 2 (CYP21A2), hydroxy-Δ-5-steroid dehydrogenase, 3β- and steroid Δ-isomerase 2 (HSD3B2), the redox partner NADPH-cytochrome P450 oxidoreductase (CPR), as well as the accessory protein cytochrome b5 (b5), a marker of the primate ZR. The rhesus ZR is mature by 3 months of age based on differentiation of the innermost zone that lacks HSD3B2, but exhibits increased b5 expression during this period. Further, the ZR develops in neonates from a previously described dense band of cells which we show expresses b5, CYP17A1, CPR, and CYP21A2 throughout maturation. The fetal zone (FZ) is distinguished from the ZR by its lack of CYP21A2, and ZR development proceeded as the FZ regressed with two important implications: neither FZ regression nor ZR maturation can be monitored independently by circulating adrenal androgens, and these events must be induced by different factors in rhesus, and likely humans. Collectively these data demonstrate that ZR development begins before birth in the rhesus, proceeding concomitantly with FZ regression post-natally, suggesting that rhesus experiences morphological adrenarche during the first three months of life.

scholarly journals Probing the Role of the Hinge Segment of Cytochrome P450 Oxidoreductase in the Interaction with Cytochrome P450

2018 ◽  
Vol 19 (12) ◽  
pp. 3914 ◽  
Author(s):  
Diana Campelo ◽  
Francisco Esteves ◽  
Bernardo Brito Palma ◽  
Bruno Costa Gomes ◽  
José Rueff ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Cytochrome B5 ◽  
Cytochrome P450s ◽  
Cytochrome P450 Reductase ◽  
Redox Partner ◽  
Redox Partners ◽  
Cytochrome P450 Oxidoreductase ◽  
Experimental Approaches ◽  
Menten Kinetic

NADPH-cytochrome P450 reductase (CPR) is the unique redox partner of microsomal cytochrome P450s (CYPs). CPR exists in a conformational equilibrium between open and closed conformations throughout its electron transfer (ET) function. Previously, we have shown that electrostatic and flexibility properties of the hinge segment of CPR are critical for ET. Three mutants of human CPR were studied (S243P, I245P and R246A) and combined with representative human drug-metabolizing CYPs (isoforms 1A2, 2A6 and 3A4). To probe the effect of these hinge mutations different experimental approaches were employed: CYP bioactivation capacity of pre-carcinogens, enzyme kinetic analysis, and effect of the ionic strength and cytochrome b5 (CYB5) on CYP activity. The hinge mutations influenced the bioactivation of pre-carcinogens, which seemed CYP isoform and substrate dependent. The deviations of Michaelis-Menten kinetic parameters uncovered tend to confirm this discrepancy, which was confirmed by CYP and hinge mutant specific salt/activity profiles. CPR/CYB5 competition experiments indicated a less important role of affinity in CPR/CYP interaction. Overall, our data suggest that the highly flexible hinge of CPR is responsible for the existence of a conformational aggregate of different open CPR conformers enabling ET-interaction with structural varied redox partners.

scholarly journals Conformational Equilibrium of NADPH–Cytochrome P450 Oxidoreductase Is Essential for Heme Oxygenase Reaction

Antioxidants ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 673
Author(s):  
Masakazu Sugishima ◽  
Junichi Taira ◽  
Tatsuya Sagara ◽  
Ryota Nakao ◽  
Hideaki Sato ◽  
...  
Keyword(s):  
Electron Transfer ◽  
Cytochrome P450 ◽  
Enzymatic Activity ◽  
Heme Oxygenase ◽  
Analytical Ultracentrifugation ◽  
Dynamic Equilibrium ◽  
P450 Oxidoreductase ◽  
Nadph Cytochrome ◽  
Redox Partner ◽  
Cytochrome P450 Oxidoreductase

Heme oxygenase (HO) catalyzes heme degradation using electrons supplied by NADPH–cytochrome P450 oxidoreductase (CPR). Electrons from NADPH flow first to FAD, then to FMN, and finally to the heme in the redox partner. Previous biophysical analyses suggest the presence of a dynamic equilibrium between the open and the closed forms of CPR. We previously demonstrated that the open-form stabilized CPR (ΔTGEE) is tightly bound to heme–HO-1, whereas the reduction in heme–HO-1 coupled with ΔTGEE is considerably slow because the distance between FAD and FMN in ΔTGEE is inappropriate for electron transfer from FAD to FMN. Here, we characterized the enzymatic activity and the reduction kinetics of HO-1 using the closed-form stabilized CPR (147CC514). Additionally, we analyzed the interaction between 147CC514 and heme–HO-1 by analytical ultracentrifugation. The results indicate that the interaction between 147CC514 and heme–HO-1 is considerably weak, and the enzymatic activity of 147CC514 is markedly weaker than that of CPR. Further, using cryo-electron microscopy, we confirmed that the crystal structure of ΔTGEE in complex with heme–HO-1 is similar to the relatively low-resolution structure of CPR complexed with heme–HO-1 in solution. We conclude that the “open–close” transition of CPR is indispensable for electron transfer from CPR to heme–HO-1.

scholarly journals Differential expression of cytochrome P450 family 4 subfamily V member 2 in human epithelial ovarian cancer.

2021 ◽  
Author(s):  
Shahan Mamoor
Keyword(s):  
Ovarian Cancer ◽  
Cytochrome P450 ◽  
Epithelial Ovarian Cancer ◽  
Fallopian Tube ◽  
Expression Profiles ◽  
Gynecologic Cancer ◽  
High Grade ◽  
Primary Tumors ◽  
Ovarian Cancers ◽  
Cytochrome P450 Family

Epithelial ovarian cancer (EOC) is the most lethal gynecologic cancer (1). We performed discovery of genes associated with epithelial ovarian cancer and of the high-grade serous ovarian cancer (HGSC) subtype, using published microarray data (2, 3) to compare global gene expression profiles of normal ovary or fallopian tube with that of primary tumors from women diagnosed with epithelial ovarian cancer or HGSC. We identified the gene encoding cytochrome P450 family 4 subfamily V member 2, CYP4V2, as among the genes whose expression was most different in epithelial ovarian cancer as compared to the normal fallopian tube. CYP4V2 expression was significantly lower in high-grade serous ovarian tumors relative to normal fallopian tube. CYP4V2 expression correlated with overall survival in patients with ovarian cancer. These data indicate that expression of CYP4V2 is perturbed in epithelial ovarian cancers broadly and in ovarian cancers of the HGSC subtype. CYP4V2 may be relevant to pathways underlying ovarian cancer initiation (transformation) or progression.

scholarly journals The Primate Adrenal Zona Reticularis is Defined by Expression of Cytochrome b5, 17α-hydroxylase/17,20-lyase Cytochrome P450 (P450c17) and NADPH-Cytochrome P450 Reductase (reductase) but not 3β-Hydroxysteroid Dehydrogenase/Δ5-4 Isomerase (3β-HSD)

1999 ◽  
Vol 84 (9) ◽  
pp. 3382-3385 ◽  
Author(s):  
Samantha Mapes ◽  
C. Jo Corbin ◽  
Alice Tarantal ◽  
Alan Conley
Keyword(s):  
Cytochrome P450 ◽  
Functional Role ◽  
Cytochrome B5 ◽  
Zona Fasciculata ◽  
Cytochrome P450 Reductase ◽  
Efficient Synthesis ◽  
Zona Reticularis ◽  
Regional Control ◽  
Biochemical Studies ◽  
Lyase Activity

Biochemical studies suggest that 17,20-lyase activity, and thus efficient synthesis of androgens by human P450c17, requires both reductase and the accessory protein cytochrome b5. Since the human and primate zona reticularis (ZR) secrete androgens, the expression of these proteins, and of 3β-HSD, was investigated by immunocytochemistry in the adrenal cortex of the mature rhesus macaque. Cytochrome b5 expression was restricted to the cells of the ZR which appeared deficient in 3β-HSD. However, both P450c17 and reductase were evident throughout the zona fasciculata. These data provide essential evidence in support of a functional role for cytochrome b5 in the regional control of 17α-hydroxylase and 17,20-lyase activities of P450c17 and thereby adrenal C19 steroid secretion by the primate adrenal gland.

scholarly journals Stereoselective Ketamine Metabolism by Genetic Variants of Cytochrome P450 CYP2B6 and Cytochrome P450 Oxidoreductase

2018 ◽  
Vol 129 (4) ◽  
pp. 756-768 ◽  
Author(s):  
Pan-Fen Wang ◽  
Alicia Neiner ◽  
Evan D. Kharasch
Keyword(s):  
Cytochrome P450 ◽  
Genetic Variants ◽  
Cytochrome B5 ◽  
Intrinsic Clearance ◽  
Cell System ◽  
Therapeutic Effects ◽  
Wild Type ◽  
P450 Oxidoreductase ◽  
Cytochrome P450 Oxidoreductase

Abstract Editor’s Perspective What We Already Know about This Topic What This Article Tells Us That Is New Background Human ketamine N-demethylation to norketamine in vitro at therapeutic concentrations is catalyzed predominantly by the cytochrome P4502B6 isoform (CYP2B6). The CYP2B6 gene is highly polymorphic. CYP2B6.6, the protein encoded by the common variant allele CYP2B6*6, exhibits diminished ketamine metabolism in vitro compared with wild-type CYP2B6.1. The gene for cytochrome P450 oxidoreductase (POR), an obligatory P450 coenzyme, is also polymorphic. This investigation evaluated ketamine metabolism by genetic variants of human CYP2B6 and POR. Methods CYP2B6 (and variants), POR (and variants), and cytochrome b5 (wild-type) were coexpressed in a cell system. All CYP2B6 variants were expressed with wild-type POR and b5. All POR variants were expressed with wild-type CYP2B6.1 and b5. Metabolism of R- and S-ketamine enantiomers, and racemic RS-ketamine to norketamine enantiomers, was determined using stereoselective high-pressure liquid chromatography–mass spectrometry. Michaelis–Menten kinetic parameters were determined. Results For ketamine enantiomers and racemate, metabolism (intrinsic clearance) was generally wild-type CYP2B6.1 > CYP2B6.4 > CYP2B6.26, CYP2B6.19, CYP2B6.17, CYP2B6.6 > CYP2B6.5, CYP2B6.7 > CYP2B6.9. CYP2B6.16 and CYP2B6.18 were essentially inactive. Activity of several CYP2B6 variants was less than half that of CYP2B6.1. CYP2B6.9 was 15 to 35% that of CYP2B6.1. The order of metabolism was wild-type POR.1 > POR.28, P228L > POR.5. CYP2B6 variants had more influence than POR variants on ketamine metabolism. Neither CYP2B6 nor POR variants affected the stereoselectivity of ketamine metabolism (S > R). Conclusions Genetic variants of CYP2B6 and P450 oxidoreductase have diminished ketamine N-demethylation activity, without affecting the stereoselectivity of metabolism. These results suggest candidate genetic polymorphisms of CYP2B6 and P450 oxidoreductase for clinical evaluation to assess consequences for ketamine pharmacokinetics, elimination, bioactivation, and therapeutic effects.

scholarly journals Cryo-EM reveals the architecture of the dimeric cytochrome P450 CYP102A1 enzyme and conformational changes required for redox partner recognition

2020 ◽  
Vol 295 (6) ◽  
pp. 1637-1645 ◽  
Author(s):  
Min Su ◽  
Sumita Chakraborty ◽  
Yoichi Osawa ◽  
Haoming Zhang
Keyword(s):  
Cytochrome P450 ◽  
Conformational Changes ◽  
Conformational Dynamics ◽  
Full Length ◽  
The Other ◽  
Redox Partner ◽  
Highly Active ◽  
Heme Domain ◽  
Fatty Acid Hydroxylation ◽  
Cytochrome P450 Family

Cytochrome P450 family 102 subfamily A member 1 (CYP102A1) is a self-sufficient flavohemeprotein and a highly active bacterial enzyme capable of fatty acid hydroxylation at a >3,000 min−1 turnover rate. The CYP102A1 architecture has been postulated to be responsible for its extraordinary catalytic prowess. However, the structure of a functional full-length CYP102A1 enzyme remains to be determined. Herein, we used a cryo-EM single-particle approach, revealing that full-length CYP102A1 forms a homodimer in which both the heme and FAD domains contact each other. The FMN domain of one monomer was located close to the heme domain of the other monomer, exhibiting a trans configuration. Moreover, full-length CYP102A1 is highly dynamic, existing in multiple conformational states, including open and closed states. In the closed state, the FMN domain closely contacts the FAD domain, whereas in the open state, one of the FMN domains rotates away from its FAD domain and traverses to the heme domain of the other monomer. This structural arrangement and conformational dynamics may facilitate rapid intraflavin and trans FMN-to-heme electron transfers (ETs). Results with a variant having a 12-amino-acid deletion in the CYP102A1 linker region, connecting the catalytic heme and the diflavin reductase domains, further highlighted the importance of conformational dynamics in the ET process. Cryo-EM revealed that the Δ12 variant homodimer is conformationally more stable and incapable of FMN-to-heme ET. We conclude that closed-to-open alternation is crucial for redox partner recognition and formation of an active ET complex for CYP102A1 catalysis.

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