The many roles of cytochrome b5

Schenkman, John B.; Jansson, Ingela

doi:10.1016/s0163-7258(02)00327-3

Cited by 406 publications

(376 citation statements)

References 124 publications

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“…The ability of CYB5A to modify the activity of CYP17A1 in the pig has been reported (Billen and Squires, 2009), and a c.-8G.T polymorphism in the CYB5A gene was associated with low androstenone levels in fat and low levels of both CYB5A mRNA and CYB5A protein (Peacock et al, 2008). The contribution of CYB5A to CYP450-mediated catalysis is complex and is dependent on the substrate and CYP450 isoform (Schenkman and Jansson, 2003). In some cases, CYB5A can modify the reaction by having an inhibitory or stimulatory effect, and in other cases it has been shown to be obligatory.…”

Section: Discussionmentioning

confidence: 99%

The roles of different porcine cytochrome P450 enzymes and cytochrome b5A in skatole metabolism

Wiercinska

Lou

Squires

2012

Animal

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Boar taint is the unfavourable odour and taste from pork fat, which results in part from the accumulation of skatole (3-methylindole, 3MI). The key enzymes in skatole metabolism are thought to be cytochrome P450 2E1 (CYP2E1) and cytochrome 2A (CYP2A); however, the cytochrome P450 (CYP450) isoform responsible for the production of the metabolite 6-hydroxy-3-methylindole (6-OH-3MI, 6-hydroxyskatole), which is thought to be involved in the clearance of skatole, has not been established conclusively. The aim of this study was to characterize the role of porcine CYP450s in skatole metabolism by expressing them individually in the human embryonic kidney HEK293-FT cell line. This system eliminates the problems of the lack of specificity of antibodies, inhibitors and substrates for CYP450 isoforms in the pig, and contributions of any other CYP450s that would be present. The results show that pig CYP1A1, CYP2A19, CYP2C33v4, CYP2C49, CYP2E1 and CYP3A and human CYP2E1 (hCYP2E1) are all capable of producing the major skatole metabolite 3-methyloxyindole (3MOI), as well as indole-3-carbinol (I3C), 5-hydroxy-3-methylindole (5-OH-3MI), 6-OH-3MI, 2-aminoacetophenone (2AAP) and 3-hydroxy-3-methyloxindole. CYP2A19 produced the highest amount of the physiologically important metabolite 6-OH-3MI, followed by porcine CYP2E1 and CYP2C49; CYP2A19 also produced more 6-OH-3MI than hCYP2E1. Co-transfection with CYB5A increased the production of skatole metabolites by some of the CYP450s, suggesting that CYB5A plays an important role in the metabolism of skatole. We also show the utility of this expression system to check the specificity of selected substrates and antibodies for porcine CYP450s. Further information regarding the abundance of different CYP450 isoforms is required to fully understand their contribution to skatole metabolism in vivo in the pig.Keywords: boar taint, skatole, 3-methylindole, cytochrome b5A, cytochrome P450 ImplicationsStudies on the metabolism and clearance of skatole, a major component of boar taint, have been hampered by the lack of specific tools to measure the activity of the cytochrome P450 (CYP450) enzymes involved. Here we have used a defined cellbased system to individually express six different isoforms of porcine CYP450 and cytochrome b5A, and measure their role in the metabolism of skatole. Our results show that CYP2A19 is the primary isoform responsible for the formation of 6-hydroxyskatole, although its importance to in vivo clearance will also be affected by its level of expression in vivo.

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Section: Discussionmentioning

confidence: 99%

The roles of different porcine cytochrome P450 enzymes and cytochrome b5A in skatole metabolism

Wiercinska

Lou

Squires

2012

Animal

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“…Moreover, cytochrome b 5 has been reported to enhance activity of certain cytochrome P450s (ref. 40). Tuning expression of the four P450s, CPR and cognate cytochrome b 5 could increase pathway efficiency.…”

Section: Rs)-norlaudanosoline (Nor) (S)-reticuline (Ret) (S)-scoulmentioning

confidence: 99%

Reconstitution of a 10-gene pathway for synthesis of the plant alkaloid dihydrosanguinarine in Saccharomyces cerevisiae

et al. 2014

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“…Since reductase and P450 form a 1:1 molar complex (3,4), those P450s not in an electron transfer complex with reductase are metabolically silent.In some cases cytochrome b 5 can transfer the second electron to particular P450s (5). Interactions between reductase and P450, reductase and cytochrome b 5 , and also P450 and cytochrome b 5 , have been described as being predominately electrostatic in nature (6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). The potential of reductase interacting with discrete populations of P450 clusters and or mobilizing P450 into smaller functional oligomers (18)(19)(20) suggests the possibility of multiple homomeric and heteromeric P450 binding sites, in addition to the residues that align P450 with redox partners.…”

mentioning

confidence: 99%

Heteromeric Complex Formation between CYP2E1 and CYP1A2: Evidence for the Involvement of Electrostatic Interactions

2006

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Mixed reconstituted systems containing CYP2B4, CYP1A2 and NADPH-cytochrome P450 reductase were previously shown to exhibit a dramatic inhibition of 7-pentoxyresorufin-Odealkylation (PROD) when compared to simple reconstituted systems containing reductase and a single P450 enzyme, results consistent with the formation of CYP1A2-CYP2B4 complexes where the reductase binds with high affinity to the CYP1A2 moiety of the complex. In this report, we provide evidence for an interaction between CYP1A2 and CYP2E1. Synergism of 7-ethoxyresorufin-Odeethylation (EROD) and PROD was observed when these P450s were combined in mixed reconstituted systems at subsaturating reductase concentrations. Higher ionic strength attenuated the synergistic stimulation of both PROD and EROD in mixed reconstituted systems, consistent with disruption of heteromeric CYP2E1-CYP1A2 complexes. The effect of ionic strength was further examined as a function of reductase concentration. At lower ionic strength, there was a significant synergistic stimulation of EROD. This synergistic stimulation diminished with increasing reductase concentration, resulting in an additive response as reductase became saturating. Interestingly, at high ionic strength, the synergism of EROD in the mixed reconstituted system was not observed. In contrast, mixed reconstituted systems containing CYP2E1 and CYP2B4 did not provide evidence for the formation of these heteromeric P450-P450 complexes. The synergistic stimulation observed with the reductase-CYP1A2-CYP2E1 mixed reconstituted system is consistent with the formation of a CYP1A2-CYP2E1 complex. Taken together with the lack of a kinetically detectable interaction between CYP2B4 and CYP2E1, and the previously reported CYP1A2-CYP2B4 interaction, these results suggest that CYP1A2 may facilitate the formation of complexes with other P450 enzymes.The cytochrome P450 (P450) superfamily of proteins is functionally diverse, and well distributed in nature. In addition to their roles in the metabolism of numerous endogenous compounds, P450s are important contributors to drug metabolism, primarily by catalyzing the insertion of an oxygen atom into a substrate molecule. NADPH cytochrome P450 reductase (reductase) is the major electron transfer partner, and is required for the majority of NADPHdependent oxidation reactions. Multiple forms of cytochrome P450 exist in the endoplasmic reticulum in a large excess over reductase (1), with the ratio of P450 to reductase dependent † These studies were supported by a US Public Health Service Research Grant from the National Institute of Enviromental Health Sciences ES004344 (WLB), and a predoctoral fellowship from the Stanley Scott Cancer Center (RWK). * Correspondence should be addressed to: Wayne L. Backes, Ph.D., Department of Pharmacology and the Stanley S. Scott Cancer Center, LSU Health Sciences Center, 533 Bolivar Street, New Orleans, La 70112, Voice 504-568-6557, FAX 504-568-6888, emailwbacke@lsuhsc.edu SUPPORTING INFORMATION AVAILABLE The supporting information includes ...

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The many roles of cytochrome b5

Cited by 406 publications

References 124 publications

The roles of different porcine cytochrome P450 enzymes and cytochrome b5A in skatole metabolism

The roles of different porcine cytochrome P450 enzymes and cytochrome b5A in skatole metabolism

Reconstitution of a 10-gene pathway for synthesis of the plant alkaloid dihydrosanguinarine in Saccharomyces cerevisiae

Heteromeric Complex Formation between CYP2E1 and CYP1A2: Evidence for the Involvement of Electrostatic Interactions

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