Isotope effects and alternative substrate reactivities for tryptophan 2,3-dioxygenase.

Leeds, Janet M.; Brown, P. J.; McGeehan, Gerard M.; Brown, Frank K.; Wiseman, Jeffrey S.

doi:10.1016/s0021-9258(17)46773-1

Cited by 57 publications

(29 citation statements)

References 34 publications

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“…The pronounced in vivo instability of 4-6-[ 18 F]FTrps and conversely the observed in vivo stability of 7-[ 18 F]FTrp could be well explained by the substrate specificity of IDO and TDO taking into account that the kynurenine pathway is responsible for the catabolism of at least 95% of Trp. Indeed, 6-FTrp is the best known substrate of TDO with V max / K m approximately 1.6-fold higher than that of Trp . 5-FTrp is also a relatively good substrate of TDO with a V max / K m value of about 70% relative to that of native tryptophan.…”

Section: Resultssupporting

confidence: 83%

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Discovery of 7-[¹⁸F]Fluorotryptophan as a Novel Positron Emission Tomography (PET) Probe for the Visualization of Tryptophan Metabolism in Vivo

et al. 2017

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Tryptophan and its metabolites are involved in different physiological and pathophysiological processes. Consequently, positron emission tomography (PET) tracers addressing tryptophan metabolic pathways should allow the detection of different pathologies like neurological disorders and cancer. Herein we report an efficient method for the preparation of fluorotryptophans labeled in different positions with F and their biological evaluation. 4-7-[F]Fluorotryptophans ([F]FTrps) were prepared according to a modified protocol of alcohol-enhanced Cu-mediated radiofluorination in 30-53% radiochemical yields. In vitro experiments demonstrated high cellular uptake of 4-7-[F]FTrps in different tumor cell lines. 4, 5-, and 6-[F]FTrps, although stable in vitro, suffered from rapid in vivo defluorination. In contrast, 7-[F]FTrp demonstrated a high in vivo stability and enabled a clear delineation of serotonergic areas and melatonin-producing pineal gland in rat brains. Moreover 7-[F]FTrp accumulated in different tumor xenografts in a chick embryo CAM model. Thus, 7-[F]FTrp represents a highly promising PET probe for imaging of Trp metabolism.

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Section: Resultssupporting

confidence: 83%

“…Furthermore, the conditions of microsomal assay were suboptimal for both heme-containing dioxygenases. For example, IDO can display its full catalytic activity only in the presence of catalase and sodium ascorbate, and TDO of Trp, sodium ascorbate, and hematin which were obviously absent in the assay medium. , …”

Section: Resultsmentioning

confidence: 99%

Discovery of 7-[¹⁸F]Fluorotryptophan as a Novel Positron Emission Tomography (PET) Probe for the Visualization of Tryptophan Metabolism in Vivo

et al. 2017

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“…To provide further experimental support for this finding, we studied the kinetic isotope effect (KIE) in cmTDO using (indole- d 5 )- l -Trp as the substrate. Our activity assays revealed a KIE value ( k H / k D ) of 0.87 ± 0.03 (Figure S3), which is much more significant than the value previously reported for rat TDO (i.e., 0.96) …”

Section: Resultsmentioning

confidence: 41%

“…Our activity assays revealed a KIE value (k H /k D ) of 0.87 ± 0.03 (Figure S3), which is much more significant than the value previously reported for rat TDO (i.e., 0.96). 63 This observed KIE represents an inverse α-secondary KIE, which is a secondary KIE (2°KIE) where the isotopic substitution occurs α to the reaction site. The α-2°KIE involves a rate difference for isotopic substitution of a bond (denoted as the "isotopic bond") that is not broken in the ratedetermining step.…”

Section: ■ Results and Discussionmentioning

confidence: 98%

Kinetic and Spectroscopic Characterization of the Catalytic Ternary Complex of Tryptophan 2,3-Dioxygenase

et al. 2020

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The first step of the kynurenine pathway for l-tryptophan (l-Trp) degradation is catalyzed by heme-dependent dioxygenases, tryptophan 2,3-dioxygenase (TDO) and indoleamine 2,3-dioxygenase. In this work, we employed stopped-flow optical absorption spectroscopy to study the kinetic behavior of the Michaelis complex of Cupriavidus metallidurans TDO (cmTDO) to improve our understanding of oxygen activation and initial oxidation of l-Trp. On the basis of the stopped-flow results, rapid freeze-quench (RFQ) experiments were performed to capture and characterize this intermediate by Mössbauer spectroscopy. By incorporating the chlorite dismutase–chlorite system to produce high concentrations of solubilized O2, we were able to capture the Michaelis complex of cmTDO in a nearly quantitative yield. The RFQ–Mössbauer results confirmed the identity of the Michaelis complex as an O2-bound ferrous species. They revealed remarkable similarities between the electronic properties of the Michaelis complex and those of the O2 adduct of myoglobin. We also found that the decay of this reactive intermediate is the rate-limiting step of the catalytic reaction. An inverse α-secondary substrate kinetic isotope effect was observed with a k H/k D of 0.87 ± 0.03 when (indole-d 5)-l-Trp was employed as the substrate. This work provides an important piece of spectroscopic evidence of the chemical identity of the Michaelis complex of bacterial TDO.

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“…Tryptophan is enzymatically oxidized at the five-position to produce 5-hydroxytryptophan and at the two- and three-positions to produce N -formylkynurenine, leading to metabolites of the serotonin and kynurenine pathways, respectively (Figure A). In principle, fluorination of the six- and seven-positions should enable normal metabolic processing of the probe; however, Wiseman and coworkers reported that tryptophan 2,3-dioxygenase oxidizes 7-F-tryptophan at a reduced rate in vitro as compared with tryptophan . Fortunately, 6-F-tryptophan has been reported to exhibit rates of oxidation at the 5- and 2,3-positions comparable to or greater than tryptophan itself. , Thus we chose to investigate 6-F-tryptophan (6-F-Trp) for our studies.…”

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confidence: 99%

Ex Vivo Analysis of Tryptophan Metabolism Using ¹⁹F NMR

Tombari

Saunders

et al. 2019

ACS Chem. Biol.

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Tryptophan, an essential amino acid, is metabolized into a variety of small molecules capable of impacting human physiology, and aberrant tryptophan metabolism has been linked to a number of diseases. There are three principal routes by which tryptophan is degraded, and thus methods for measuring metabolic flux through these pathways can be used to understand the factors that perturb tryptophan metabolism and potentially to measure disease biomarkers. Here, we describe a method utilizing 6fluorotryptophan as a probe for detecting tryptophan metabolites in ex vivo tissue samples via 19 F nuclear magnetic resonance. As a proof of concept, we demonstrate that 6-fluorotryptophan can be used to measure changes in tryptophan metabolism resulting from antibiotic-induced changes in gut microbiota composition. Taken together, we describe a general strategy for monitoring amino acid metabolism using 19 F nuclear magnetic resonance that is operationally simple and does not require chromatographic separation of metabolites.

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Isotope effects and alternative substrate reactivities for tryptophan 2,3-dioxygenase.

Cited by 57 publications

References 34 publications

Discovery of 7-[¹⁸F]Fluorotryptophan as a Novel Positron Emission Tomography (PET) Probe for the Visualization of Tryptophan Metabolism in Vivo

Discovery of 7-[¹⁸F]Fluorotryptophan as a Novel Positron Emission Tomography (PET) Probe for the Visualization of Tryptophan Metabolism in Vivo

Kinetic and Spectroscopic Characterization of the Catalytic Ternary Complex of Tryptophan 2,3-Dioxygenase

Ex Vivo Analysis of Tryptophan Metabolism Using ¹⁹F NMR

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Isotope effects and alternative substrate reactivities for tryptophan 2,3-dioxygenase.

Cited by 57 publications

References 34 publications

Discovery of 7-[18F]Fluorotryptophan as a Novel Positron Emission Tomography (PET) Probe for the Visualization of Tryptophan Metabolism in Vivo

Discovery of 7-[18F]Fluorotryptophan as a Novel Positron Emission Tomography (PET) Probe for the Visualization of Tryptophan Metabolism in Vivo

Kinetic and Spectroscopic Characterization of the Catalytic Ternary Complex of Tryptophan 2,3-Dioxygenase

Ex Vivo Analysis of Tryptophan Metabolism Using 19F NMR

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Discovery of 7-[¹⁸F]Fluorotryptophan as a Novel Positron Emission Tomography (PET) Probe for the Visualization of Tryptophan Metabolism in Vivo

Discovery of 7-[¹⁸F]Fluorotryptophan as a Novel Positron Emission Tomography (PET) Probe for the Visualization of Tryptophan Metabolism in Vivo

Ex Vivo Analysis of Tryptophan Metabolism Using ¹⁹F NMR