Alkylation of the prosthetic heme in cytochrome P-450 during oxidative metabolism of the sedative-hypnotic ethchlorvynol

Pr, Ortiz de Montellano; Hs, Beilan; Jm, Mathews

doi:10.1021/jm00352a015

Cited by 25 publications

(10 citation statements)

References 6 publications

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“…However, whether the oxygen has been added to the internal or terminal carbon cannot be ascertained from the MS/MS spectrum. Ortiz de Montellano et al, (1982) have previously reported that if the oxygen atom is added to the internal carbon, the heme will be alkylated, and if the oxygen is added to the terminal carbon, the protein will be modified (Ortiz de Montellano and Komives, 1985;CaJacob et al, 1988;Chan et al, 1993). Because a protein adduct was identified in this case, the structure proposed for the GSH conjugate has the oxygen inserted into the terminal carbon as shown in Fig.…”

Section: Lc/ms/ms Analysis Of Gsh Conjugates Of Bpa and Bmpmentioning

confidence: 89%

“…Ortiz de Montellano et al (1982) have shown the formation of three metal-free regioisomers as a result of prosthetic heme alkylation after administration of ethchlorvynol to phenobarbital-treated rats. The first two isomers, which eluted very close together by HPLC, are identified as structures with the N-alkyl group on the vinyl-substituted pyrrole rings (A and B).…”

Section: Discussionmentioning

confidence: 99%

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Mechanism-Based Inactivation of CYP2B1 and Its F-Helix Mutant by Twotert-Butyl Acetylenic Compounds: Covalent Modification of Prosthetic Heme Versus Apoprotein

Lin¹,

Zhang²,

Noon³

et al. 2009

J Pharmacol Exp Ther

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The mechanism-based inactivation of cytochrome CYP2B1 [wild type (WT)] and its Thr205 to Ala mutant (T205A) by tertbutylphenylacetylene (BPA) and tert-butyl 1-methyl-2-propynyl ether (BMP) in the reconstituted system was investigated. The inactivation of WT by BPA exhibited a k inact /K I value of 1343 min Ϫ1 mM Ϫ1 and a partition ratio of 1. The inactivation of WT by BMP exhibited a k inact /K I value of 33 min Ϫ1 mM Ϫ1 and a partition ratio of 10. Liquid chromatography/tandem mass spectrometry analysis (LC/MS/MS) of the WT revealed 1) inactivation by BPA resulted in the formation of a protein adduct with a mass increase equivalent to the mass of BPA plus one oxygen atom, and 2) inactivation by BMP resulted in the formation of multiple heme adducts that all exhibited a mass increase equivalent to BMP plus one oxygen atom. LC/MS/MS analysis indicated the formation of glutathione (GSH) conjugates by the reaction of GSH with the ethynyl moiety of BMP or BPA with the oxygen being added to the internal or terminal carbon. For the inactivation of T205A by BPA and BMP, the k inact /K I values were suppressed by 100-and 4-fold, respectively, and the partition ratios were increased 9-and 3.5-fold, respectively. Only one major heme adduct was detected following the inactivation of the T205A by BMP. These results show that the Thr205 in the F-helix plays an important role in the efficiency of the mechanism-based inactivation of CYP2B1 by BPA and BMP. Homology modeling and substrate docking studies were presented to facilitate the interpretation of the experimental results.

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Section: Lc/ms/ms Analysis Of Gsh Conjugates Of Bpa and Bmpmentioning

confidence: 89%

Section: Discussionmentioning

confidence: 99%

Mechanism-Based Inactivation of CYP2B1 and Its F-Helix Mutant by Twotert-Butyl Acetylenic Compounds: Covalent Modification of Prosthetic Heme Versus Apoprotein

Lin¹,

Zhang²,

Noon³

et al. 2009

J Pharmacol Exp Ther

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“…The iron-porphyrin complexes have been unambiguously identified as o-iron(III)-CH3 and o-iron(III)-CsHS complexes (Scheme 1) [15]. Moreover, evidence has been provided for similar o-phenyl [2,17] and o-methyl (our unpublished results) structures for the corresponding hemoglobin or myoglobin complexes. This analogy leads us to propose a cytochrome P450-Fe(III)-R structure with R=CH, or C,H,, for the complexes derived from reactions of microsomal cytochrome P450 with methylhydrazine or phenylhydrazine derivatives, which exhibit a Soret peak around 480nm.…”

Section: Nature Of' the Cytochrome P450 Complexes Exhibiting A Soret mentioning

confidence: 92%

Reaction of Monosubstituted Hydrazines and Diazenes with Rat‐Liver Cytochrome P450

Battioni

Mahy

Delaforge

et al. 1983

European Journal of Biochemistry

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The alkyldiazenes RN = NH (R = CH3 or CzHs) react with reduced microsomal cytochrome P450 leading to complexes exhibiting a Soret peak at 446 nm. Upon oxidation of the [cytochrome P450-Fe(II)(CH3N = NH)] complex with limited amounts of dioxygen, a new complex characterized by a Soret peak at 486 nm is formed. The latter complex was also formed upon slow reaction of methyldiazene with microsomal cytochrome P450-Fe(III) or in situ oxidation of methylhydrazine by limited amounts of O2 or ferricyanide. This complex is rapidly destroyed by 0 2 or ferricyanide in excess and more slowly by excess dithionite in the presence of CO.Reactions of ethyldiazene or benzyldiazene with cytochrome P450-Fe(I 11) afforded similar complexes characterized by Soret peaks around 480 nm.These results, when compared to those recently described on reactions of monosubstituted hydrazines RNHNH2 and diazenes R N = N H with hemoglobin and iron-porphyrins, are consistent with a [cytochrome P450-Fe(II)(RN = NH)] structure for the 446-nm-absorbing complexes and a a-alkyl cytochrome P450-Fe(III)-R structure for the complexes characterized by a Soret peak around 480 nm. They also suggest a a-cytochrome P450-Fe(III)-Ph structure for the complex derived from phenylhydrazine oxidation, recently described in the literature. Finally, they provide the first evidence that cytochrome P450-Fe(III)-R complexes are formed upon microsomal oxidation of alkyl or phenylhydrazines. PFe"The dioxygen-dependent oxidation of monosubstituted hydrazines in the presence of iron-porphyrins has been recently studied [I51 and the successive formation of two ironligand complexes in the reaction has been established (Scheme 1). The first complex derives from the binding of the diazene RN = NH, the two-electron oxidation product of the starting hydrazine RNHNH2, to the iron(I1)-porphyrin. The second complex is the o-alkyl (or a-aryl) porphyriniron(II1)-R complex which is formed by a one-electron oxidation of the didzene-iron(1I) complex. The iron(II1)-methyl

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“…Most importantly, these losses are associated with accumulation in the liver (or in incubations with tissue microsomes) of a green, red-fluorescing pigment (Tables 1 and 2). Isolation and characterization by UV-visible, mass spectrometric and NMR methods of several of these pigments has shown that in all of them the acetylene is covalently attached to one of the nitrogen atoms of protoporphyrin IX, the framework of the prosthetic heme group of the enzyme (e.g., Ortiz de Montellano et al 1979, 1982a, 1982bKunze 1980b, 1981b;Kunze et al 1983). In these adducts, the terminal carbon of the acetylene is attached to a porphyrin ring nitrogen atom and the internal carbon of the acetylene is present as a carbonyl group, as shown for the adduct obtained with propyne in Figure 15.…”

Section: Prosthetic Heme Alkylation By Acetylene and Monosubstituted mentioning

confidence: 99%

“…The four pyrrole rings of the porphyrin are labeled A-D. In this instance, the covalent bond was formed exclusively with pyrrole ring A, but other pyrrole rings can be alkylated in other adducts (e.g., Ortiz de Montellano et al 1982a). Schematic mechanism for autocatalytic alkylation of the heme prosthetic group of cytochrome P450 in the oxidation of a terminal acetylene.…”

Section: Specificitymentioning

confidence: 99%

Acetylenes: cytochrome P450 oxidation and mechanism-based enzyme inactivation

Montellano

2019

Drug Metabolism Reviews

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The oxidation of carbon-carbon triple bonds by cytochrome P450 produces ketene metabolites that are hydrolyzed to acetic acid derivatives or are trapped by nucleophiles. In the special case of 17αethynyl sterols, D-ring expansion and de-ethynylation have been observed as competing pathways. The oxidation of acetylenic groups is also associated with mechanism-based inactivation of cytochrome P450 enzymes. One mechanism for this inactivation is reaction of the ketene metabolite with cytochrome P450 residues essential for substrate binding or catalysis. However, in the case of monosubstituted acetylenes, inactivation can also occur by addition of the oxidized acetylenic function to a nitrogen of the heme prosthetic group. This addition reaction is not mediated by the ketene metabolite, but rather occurs during oxygen transfer to the triple bond. In some instances, a detectable intermediate is formed that is most consistent with a ketocarbene-iron heme complex. This complex can progress to the N-alkylated heme or revert back to the unmodified enzyme. The ketocarbene complex may intervene in the formation of all the N-alkyl heme adducts, but is normally too unstable to be detected.

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Alkylation of the prosthetic heme in cytochrome P-450 during oxidative metabolism of the sedative-hypnotic ethchlorvynol

Cited by 25 publications

References 6 publications

Mechanism-Based Inactivation of CYP2B1 and Its F-Helix Mutant by Twotert-Butyl Acetylenic Compounds: Covalent Modification of Prosthetic Heme Versus Apoprotein

Mechanism-Based Inactivation of CYP2B1 and Its F-Helix Mutant by Twotert-Butyl Acetylenic Compounds: Covalent Modification of Prosthetic Heme Versus Apoprotein

Reaction of Monosubstituted Hydrazines and Diazenes with Rat‐Liver Cytochrome P450

Acetylenes: cytochrome P450 oxidation and mechanism-based enzyme inactivation

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