Characterization of the MethionineS-Oxidase Activity of Rat Liver and Kidney Microsomes: Immunochemical and Kinetic Evidence for FMO3 Being the Major Catalyst

Krause, Renee J.; Ripp, Sharon L.; Sausen, Peter J.; Overby, Lila H.; Philpot, Richard M.; Elfarra, Adnan A.

doi:10.1006/abbi.1996.0370

Cited by 34 publications

(32 citation statements)

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“…Although we did not determine kinetic values with cDNA-expressed human FMOs, we previously have shown that for many substrates such as Lmethionine, S-allyl-L-cysteine, and S-benzyl-L-cysteine, kinetic values obtained with either purified rat liver FMOs or cDNAexpressed rabbit FMOs were similar to the corresponding values obtained with cDNAexpressed human FMOs (13,15,29). With SeMet, the V max values obtained with purified rat liver FMO1 and FMO3 (1200 and 280 nmol/mg/min, respectively) are similar to the corresponding values (1266 and 261 nmol/mg/min, respectively) with L-methionine (14). Interestingly, with both SeMet and L-methionine, the V max /K m ratio is much higher with rat liver FMO3 than with rat liver FMO1.…”

Section: Discussionsupporting

confidence: 62%

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Oxidative Metabolism of Seleno-l-methionine to l-Methionine Selenoxide by Flavin-Containing Monooxygenases

Krause

Glocke

Sicuri

et al. 2006

Chem. Res. Toxicol.

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The roles of flavin-containing monooxygenases (FMOs) in the oxidation of seleno-L-methionine (SeMet) to L-methionine selenoxide (MetSeO) were investigated using cDNA-expressed human FMOs, purified rat liver FMOs, and rat liver microsomes. MetSeO and the N-2,4-dinitrophenylderivatives of SeMet and MetSeO were synthesized and characterized by 1 H-NMR and ESI/MS. These reference compounds were then used to develop a sensitive HPLC assay to monitor SeMet oxidation to MetSeO. Formation of MetSeO in rat liver microsomes was time-, protein concentration-, SeMet concentration-, and NADPH-dependent. The microsomal activity exhibited a SeMet K m value (mean ±S.D.; n=4) of 0.91 ± 0.29 mM and a V max value of 44 ± 8.0 nmol MetSeO/ mg protein/min. Inclusion of 1-benzylimidazole, superoxide dismutase or deferoxamine caused no inhibition of the rat liver microsomal activity. Because these results suggested the involvement of FMOs in the oxidation of SeMet in rat liver microsomes, formation of MetSeO was also examined using cDNA-expressed human and purified rat FMOs. The results showed that both rat and human FMO1 and FMO3 but not FMO5 can catalyze the reaction. The SeMet kinetic constants were obtained with purified rat liver FMO3 (K m = 0.11 mM, V max = 280 nmol/mg protein/min) and rat liver FMO1 (K m = 7.8 mM, V max = 1200 nmol/mg protein/min). Because SeMet has anti-cancer, chemopreventive, and toxic properties, the kinetic results suggest FMO3 is likely to play a role in the biological activities of SeMet at low exposure conditions.

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Section: Discussionsupporting

confidence: 62%

“…After these metabolic reactions were terminated using an equal volume of ice-cold ethanol, samples were derivatized with 1-fluoro-2,4-dinitrobenzene and analyzed for methionine sulfoxide formation as previously described (14).…”

Section: Cautionmentioning

confidence: 99%

Oxidative Metabolism of Seleno-l-methionine to l-Methionine Selenoxide by Flavin-Containing Monooxygenases

Krause

Glocke

Sicuri

et al. 2006

Chem. Res. Toxicol.

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“…Many compounds containing a negative charge are excluded entirely or are poor substrates with exceptions such as sulindac sulfide and lipoic acid (Taylor & Ziegler, 1987;Hvattum et al, 1991;Attar et al, 2003). Zwitterions and compounds with more than 1 positive charge are typically not substrates, although some, like methionine, can be oxygenated, but the apparent K m is above physiological levels (Park et al, 1992(Park et al, , 1994Duescher et al, 1994;Elfarra, 1995;Krause et al, 1996, Ripp et al, 1997, 1999. Diamines, such as cadaverine and spermidine, are not oxygenated by FMO.…”

Section: Substrate Specificity Of Enzyme-restricted Substrate Access mentioning

confidence: 99%

Mammalian flavin-containing monooxygenases: structure/function, genetic polymorphisms and role in drug metabolism

Krueger

Williams

2005

Pharmacology & Therapeutics

515

444

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Flavin-containing monooxygenase (FMO) oxygenates drugs and xenobiotics containing a "softnucleophile", usually nitrogen or sulfur. FMO, like cytochrome P450 (CYP), is a monooxygenase, utilizing the reducing equivalents of NADPH to reduce 1 atom of molecular oxygen to water, while the other atom is used to oxidize the substrate. FMO and CYP also exhibit similar tissue and cellular location, molecular weight, substrate specificity, and exist as multiple enzymes under developmental control. The human FMO functional gene family is much smaller (5 families each with a single member) than CYP. FMO does not require a reductase to transfer electrons from NADPH and the catalytic cycle of the 2 monooxygenases is strikingly different. Another distinction is the lack of induction of FMOs by xenobiotics.In general, CYP is the major contributor to oxidative xenobiotic metabolism. However, FMO activity may be of significance in a number of cases and should not be overlooked. FMO and CYP have overlapping substrate specificities, but often yield distinct metabolites with potentially significant toxicological/pharmacological consequences.The physiological function(s) of FMO are poorly understood. Three of the 5 expressed human FMO genes, FMO1, FMO2 and FMO3, exhibit genetic polymorphisms. The most studied of these is FMO3 (adult human liver) in which mutant alleles contribute to the disease known as trimethylaminuria. The consequences of these FMO genetic polymorphisms in drug metabolism and human health are areas of research requiring further exploration.

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“…The V/K value for FMO4 was not determined. These data indicated that FMO3 S-oxidation of Met proceeds with the highest affinity and greatest diastereomeric selectivity among FMO1-5.Stereoselective formation of Met-d-O was also detected in rabbit and rat liver microsomes incubated with Met, and exhibited K m and V/K values similar to those of cDNA-expressed FMO3 (Duescher et al, 1994;Krause et al, 1996). The latter results provided evidence for FMO3 being the primary isoform involved in Met S-oxidation in rabbit and rat liver.…”

mentioning

confidence: 91%

In Vivo Metabolism of l-Methionine in Mice: Evidence for Stereoselective Formation of Methionine-d-Sulfoxide and Quantitation of Other Major Metabolites

Dever¹,

Elfarra²

2006

Drug Metabolism and Disposition

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Flavin-containing monooxygenases (FMOs) are microsomal enzymes that catalyze the NADPH-and O 2 -dependent oxidation of heteroatoms (nitrogen, sulfur, phosphorus) present in the chemical structure of a variety of drugs and xenobiotics. Five functional forms (FMO1-5) have been characterized to date (Ziegler, 2002). In humans, FMO1 is the primary isoform expressed in neonate liver, but a switch occurs shortly after birth to FMO3, the primary isoform expressed in adult human liver (Koukouritaki et al., 2002). In mice, females, but not males, also express FMO3 in the liver (Falls et al., 1995;Ripp et al., 1999b). This sex-specific expression of FMO3 in mice makes them an attractive model for studying the role of FMO3 in human drug metabolism and disease.Our laboratory first identified Met as a substrate for cDNA-expressed rabbit FMO1-3 with K m values of 48.0, 29.9, and 6.5 mM, respectively (Duescher et al., 1994). The V/K values (0.9, 1.7, and 6.1 for FMO1-3, respectively) also suggested that FMO3 was the most efficient Met S-oxidizer of these three FMO isoforms. FMO3 Soxidation of Met was highly stereoselective, forming 8.4 times more methionine-d-sulfoxide (Met-d-O) than methionine-l-sulfoxide (Metl-O), whereas the d:l diastereomeric ratios for FMO1 and FMO2 were 1.5:1 and 0.7:1, respectively; FMO5 S-oxidation of Met was not detected. Recombinant human FMO3 exhibited a K m value of 3.7 mM with a V/K value of 4.6 and resulted in stereoselective formation of (Ripp et al., 1999b). Recombinant human FMO4 S-oxidation of Met exhibited a K m value greater than 10 mM with only 30% of the total sulfoxide formed being Met-d-O (Ripp et al., 1999a). The V/K value for FMO4 was not determined. These data indicated that FMO3 S-oxidation of Met proceeds with the highest affinity and greatest diastereomeric selectivity among FMO1-5.Stereoselective formation of Met-d-O was also detected in rabbit and rat liver microsomes incubated with Met, and exhibited K m and V/K values similar to those of cDNA-expressed FMO3 (Duescher et al., 1994;Krause et al., 1996). The latter results provided evidence for FMO3 being the primary isoform involved in Met S-oxidation in rabbit and rat liver. Additional experiments also provided evidence that FMOs, but not cytochrome P450s, peroxidases, or reactive oxygen species, mediated the Met S-oxidase activity (Krause et al., 1996).

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Characterization of the MethionineS-Oxidase Activity of Rat Liver and Kidney Microsomes: Immunochemical and Kinetic Evidence for FMO3 Being the Major Catalyst

Cited by 34 publications

References 0 publications

Oxidative Metabolism of Seleno-l-methionine to l-Methionine Selenoxide by Flavin-Containing Monooxygenases

Oxidative Metabolism of Seleno-l-methionine to l-Methionine Selenoxide by Flavin-Containing Monooxygenases

Mammalian flavin-containing monooxygenases: structure/function, genetic polymorphisms and role in drug metabolism

In Vivo Metabolism of l-Methionine in Mice: Evidence for Stereoselective Formation of Methionine-d-Sulfoxide and Quantitation of Other Major Metabolites

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