Functional analysis of the cytochrome P450 monooxygenase Gene bcbot1 of Botrytis cinerea indicates. That botrydial is a strain-specific virulence factor

The mechanism-based inactivation of the cytochrome P450 (P450) dependent monooxygenase system was studied in vivo in liver, lung, and kidney of untreated, phenobarbital-treated, and β-naphthoflavone-treated guinea pigs 24 h after administration of 1-aminobenzotriazole (1 – 100 mg/kg, i.p.). Microsomal isozyme-selective or -specific monooxygenase activities were inhibited in a dose-dependent manner in all three organs. In the liver of untreated and phenobarbital-treated animals, 7-pentoxyresorufin O-depentylation (catalyzed primarily by P450 2Bx, an orthologue of rabbit P450 2B4/rat 2B1) was inhibited more than 7-ethoxyresorufin O-deethylation (P450 1A1), 4-aminobiphenyl N-hydroxylation (P450 1A2), erythromycin N-demefhylation (P450 3A), or benzphetamine N-demethylation; in β-naphthoflavone-treated animals, 4-aminobiphenyl N-hydroxylation activity was preferentially inhibited. In lung, the order of inactivation of monooxygenase activities was 4-aminobiphenyl N-hydroxylation (4Bx, the orthologue of rabbit 4B1) > 7-pentoxyresorufin O-depentylation activity (2Bx) > 7-ethoxyresorufin O-deethylation (1A1; for example 72, 53, and 29% inactivation, respectively, in phenobarbital-treated animals at 100 mg/kg). In all three tissues the loss in spectrally assayed P450 content corresponds quite well to the inhibition of monooxygenase activities. Thus, these studies show that 1-aminobenzotriazole is an effective inactivator of the pulmonary, hepatic, and renal P450 systems in guinea pigs following i.p. administration, and that P450 1A2 (liver) and P450 4Bx (lung), isozymes efficient for the oxidation of primary aromatic amines, are preferentially inactivated.Key words: cytochrome P450, 1-aminobenzotriazole, mechanism-based inhibition, lung, liver.

Phenobarbital

Hydroxylation

Benzphetamine

10.1139/y92-231

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Citations (33)

Evaluation of the stereoselective biotransformation of permethrin in human liver microsomes: Contributions of cytochrome P450 monooxygenases to the formation of estrogenic metabolites

Toxicology Letters (2014)

Ramón Lavado Jiwen Li John M. Rimoldi Daniel Schlenk

Biotransformation

Hydroxylation

Microsoma

10.1016/j.toxlet.2014.02.005

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Citations (22)

[Expression and characterization of a novel cytochrome P450 enzyme from Variovorax paradoxus S110].

PubMed (2020)

Chenxing Li Xiaodong Hou Baodang Guo Y. Manjula Rao

Cytochrome P450 monooxygenases as powerful biocatalysts catalyze a wide range of chemical reactions to facilitate exogenous substances metabolism and biosynthesis of natural products. In order to explore new catalytic reactions and increase the number of P450 biocatalysts used in synthetic biology, a new self-sufficient cytochrome P450 monooxygenase (P450(VpMO)), belongs to CYP116B class, was mined from Variovorax paradoxus S110 genome and expressed in Escherichia coli. Based on characterization of the enzymatic properties, it shows that the optimal pH and temperature for P450(VpMO) reaction activity are 8.0 and 45 °C, respectively. P450(VpMO) is relatively stable at temperatures below 35 °C. The Km and kcat of P450(VpMO) toward 4-Methoxyacetophenone are 0.458 mmol/L and 2.438 min⁻¹, respectively. Importantly, P450(VpMO) was able to catalyze the demethylation reaction for a range of substrates containing methoxy group. Its demethylation reactivity is reasonably better than other P450s belongs to CYP116B class, particularly, for 4-methoxyacetophenone with a great conversion efficiency at 91%, showing that P450(VpMO) could be used as a great biocatalyst candidate for further analysis.细胞色素P450 单加氧酶 (Cytochrome P450 monooxygenases) 是一种广谱催化剂，可以催化多种类型反应而参与生物体外源物质代谢与天然产物的合成。为丰富P450 作为合成生物学的酶元件库，并探索新型催化反应，利用生物信息学手段从争论贪噬菌Variovorax paradoxus S110 中挖掘出一种新型电子自供体细胞色素P450(VpMO) 单加氧酶，属于CYP116B 家族，它可以在大肠杆菌Escherichia coli 异源可溶表达。酶学性质研究表明P450(VpMO) 最适pH 和最适温度分别为8.0 和45 ℃，并且在温度低于35 ℃时具有良好的稳定性，Km值为0.458 mmol/L，kcat 为2.438 min⁻¹；重要的是重组P450(VpMO) 可以催化一系列包含污染物的含甲氧基底物进行脱甲基反应，其中对4-甲氧基苯乙酮的脱甲基反应转化率高达91%。相比于其他CYP116B 家族的P450 酶，P450(VpMO) 表现出较强的酶活性，这为后期进一步研究P450(VpMO) 提供了基础。.

Enzyme Kinetics

Demethylation

Biocatalysis

10.13345/j.cjb.190533

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Determination of the Dominant Arachidonic Acid Cytochrome P450 Monooxygenases in Rat Heart, Lung, Kidney, and Liver: Protein Expression and Metabolite Kinetics

The AAPS Journal (2012)

Ahmed A. El-Sherbeni Mona E. Aboutabl Beshay N. Zordoky Anwar Anwar‐Mohamed Ayman O.S. El‐Kadi

Hydroxylation

Hydroxyeicosatetraenoic acid

10.1208/s12248-012-9425-7

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Citations (43)

Pesticide-metabolizing enzymes

Toxicology Letters (1995)

Ernest Hodgson Randy L. Rose Doug‐Young Ryu Greg Falls Bonnie L. Blake

Flavin-containing monooxygenase

Xenobiotic

10.1016/0378-4274(95)03469-2

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Citations (59)

Catalysis with CytochromeP450 Monooxygenases

Vlada B. Urlacher

Abstract Cytochromes P450 belong to the ever‐growing superfamily of heme‐containing monooxygenases with currently more than 7700 gene sequences found in all domains of life. Cytochrome P450 enzymes play a central role in drug metabolism and have been shown to be involved in the biosynthesis of important natural compounds. These enzymes convert a vast range of substrates and catalyze diverse reactions. Their properties have been used for drug development, bioremediation and the synthesis of fine chemicals. Significant progress has been made over the past decade in engineering P450 monooxygenases with widely altered substrate specificities, substantially increased activity and enhanced stability. Furthermore, several approaches for the replacement or regeneration of the cofactor NAD(P)H have been developed, allowing the more cost‐effective use of P450 enzymes. In this chapter, we focus on the recent progress in applying P450 monooxygenases as biocatalysts. The P450 engineering issue will also be discussed.

Protein Engineering

10.1002/9783527628698.hgc025

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Manipulating virulence factor availability can have complex consequences for infections

bioRxiv (Cold Spring Harbor Laboratory) (2016)

Michael Weigert Adin Ross‐Gillespie Anne Leinweber Gabriella Pessi Sam P. Brown

Abstract Given the rise of bacterial resistance against antibiotics, we urgently need alternative strategies to fight infections. Some propose we should disarm rather than kill bacteria, through targeted disruption of their virulence factors. It is assumed that this approach (i) induces weak selection for resistance because it should only minimally impact bacterial fitness, and (ii) is specific, only interfering with the virulence factor in question. Given that pathogenicity emerges from complex interactions between pathogens, hosts, and their environment, such assumptions may be unrealistic. To address this issue in a test case, we conducted experiments with the opportunistic human pathogen Pseudomonas aeruginosa , where we manipulated the availability of a virulence factor, the iron-scavenging pyoverdine, within the insect host Galleria mellonella . We observed that pyoverdine availability was not stringently predictive of virulence, and affected bacterial fitness in non-linear ways. We show that this complexity could partly arise because pyoverdine availability affects host responses and alters the expression of regulatorily linked virulence factors. Our results reveal that virulence-factor manipulation feeds back on pathogen and host behavior, which in turn affects virulence. Our findings highlight that realizing effective and evolutionarily robust anti-virulence therapies will ultimately require deeper engagement with the intrinsic complexity of host-pathogen systems.

Virulence factor

Galleria mellonella

Pyoverdine

Biodefense

Human pathogen

10.1101/062570

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Citations (1)