Exploring the mechanism of tryptophan 2,3-dioxygenase

The haem proteins TDO (tryptophan 2,3-dioxygenase) and IDO (indoleamine 2,3-dioxygenase) are specific and powerful oxidation catalysts that insert one molecule of dioxygen into L-tryptophan in the first and rate-limiting step in the kynurenine pathway. Recent crystallographic and biochemical analyses of TDO and IDO have greatly aided our understanding of the mechanisms employed by these enzymes in the binding and activation of dioxygen and tryptophan. In the present paper, we briefly discuss the function, structure and possible catalytic mechanism of these enzymes.

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Oxidation of L-tryptophan in biology: a comparison between tryptophan 2,3-dioxygenase and indoleamine 2,3-dioxygenase

Biochemical Society Transactions ◽

10.1042/bst0370408 ◽

2009 ◽

Vol 37 (2) ◽

pp. 408-412 ◽

Cited By ~ 54

Author(s):

Sara A. Rafice ◽

Nishma Chauhan ◽

Igor Efimov ◽

Jaswir Basran ◽

Emma Lloyd Raven

Keyword(s):

Structural Information ◽

Recombinant Expression ◽

Kynurenine Pathway ◽

Expression Systems ◽

Indoleamine 2,3 Dioxygenase ◽

Rate Limiting Step ◽

Limiting Step ◽

The Family ◽

And Function ◽

Rate Limiting

The family of haem dioxygenases catalyse the initial oxidative cleavage of L-tryptophan to N-formylkynurenine, which is the first, rate-limiting, step in the L-kynurenine pathway. In the present paper, we discuss and compare structure and function across the family of haem dioxygenases by focusing on TDO (tryptophan 2,3-dioxygenase) and IDO (indoleamine 2,3-dioxygenase), including a review of recent structural information for both enzymes. The present paper describes how the recent development of recombinant expression systems has informed our more detailed understanding of the substrate binding, catalytic activity and mechanistic properties of these haem dioxygenases.

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Substrate-assisted mechanism of catalytic hydrolysis of misaminoacylated tRNA required for protein synthesis fidelity

Biochemical Journal ◽

10.1042/bcj20180910 ◽

2019 ◽

Vol 476 (4) ◽

pp. 719-732 ◽

Cited By ~ 1

Author(s):

Mykola M. Ilchenko ◽

Mariia Yu. Rybak ◽

Alex V. Rayevsky ◽

Oksana P. Kovalenko ◽

Igor Ya. Dubey ◽

...

Keyword(s):

Amino Acids ◽

Catalytic Mechanism ◽

Thermus Thermophilus ◽

Extensive Study ◽

Water Molecules ◽

Amino Acid Residues ◽

Rate Limiting Step ◽

Limiting Step ◽

Mechanism Of Hydrolysis ◽

Rate Limiting

Abstract d-aminoacyl-tRNA-deacylase (DTD) prevents the incorporation of d-amino acids into proteins during translation by hydrolyzing the ester bond between mistakenly attached amino acids and tRNAs. Despite extensive study of this proofreading enzyme, the precise catalytic mechanism remains unknown. Here, a combination of biochemical and computational investigations has enabled the discovery of a new substrate-assisted mechanism of d-Tyr-tRNATyr hydrolysis by Thermus thermophilus DTD. Several functional elements of the substrate, misacylated tRNA, participate in the catalysis. During the hydrolytic reaction, the 2′-OH group of the А76 residue of d-Tyr-tRNATyr forms a hydrogen bond with a carbonyl group of the tyrosine residue, stabilizing the transition-state intermediate. Two water molecules participate in this reaction, attacking and assisting ones, resulting in a significant decrease in the activation energy of the rate-limiting step. The amino group of the d-Tyr aminoacyl moiety is unprotonated and serves as a general base, abstracting the proton from the assisting water molecule and forming a more nucleophilic ester-attacking species. Quantum chemical methodology was used to investigate the mechanism of hydrolysis. The DFT-calculated deacylation reaction is in full agreement with the experimental data. The Gibbs activation energies for the first and second steps were 10.52 and 1.05 kcal/mol, respectively, highlighting that the first step of the hydrolysis process is the rate-limiting step. Several amino acid residues of the enzyme participate in the coordination of the substrate and water molecules. Thus, the present work provides new insights into the proofreading details of misacylated tRNAs and can be extended to other systems important for translation fidelity.

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Serum Indoleamine 2,3-Dioxygenase Activity Predicts Prognosis of Pulmonary Tuberculosis

Clinical and Vaccine Immunology ◽

10.1128/cvi.05402-11 ◽

2012 ◽

Vol 19 (3) ◽

pp. 436-442 ◽

Cited By ~ 55

Author(s):

Yuzo Suzuki ◽

Takafumi Suda ◽

Kazuhiro Asada ◽

Seiichi Miwa ◽

Masako Suzuki ◽

...

Keyword(s):

Poor Prognosis ◽

Roc Analysis ◽

Operating Characteristic ◽

Area Under The Curve ◽

Major Health Problem ◽

Major Health ◽

Indoleamine 2,3 Dioxygenase ◽

Rate Limiting Step ◽

Limiting Step ◽

Rate Limiting

ABSTRACTTuberculosis (TB) continues to be a major health problem, and there are few biomarkers for predicting prognosis. Indoleamine 2,3-dioxygenase (IDO), a potent immunoregulatory molecule, catalyzes the rate-limiting step of tryptophan (Trp) degradation in the kynurenine (Kyn) pathway. An increase in IDO activity determined by the serum Trp/Kyn ratio has been shown to be associated with poor prognosis in cancers and bacteremia. In TB, however, there are no studies measuring serum IDO activity to determine its clinical significance. We evaluated serum IDO activity with 174 pulmonary TB (PTB) patients and 85 controls, using liquid chromatography/electrospray ionization tandem mass spectrometry. IDO activity was estimated by calculating the serum Kyn-to-Trp ratio. PTB patients had significantly higher Kyn concentrations and IDO activity and significantly lower Trp concentrations (P< 0.0001,P< 0.0001, andP< 0.0001, respectively) than the controls. Of 174 PTB patients, 39 (22.4%) died. The patients who died had significantly higher concentrations of Kyn and significantly lower Trp concentrations, resulting in significantly higher IDO activity (P< 0.0001,P< 0.0001, andP< 0.0001, respectively). In a receiver operating characteristic (ROC) analysis, serum IDO activity had the highest area under the curve (0.850), and this activity was an independent prognostic factor in multivariate analysis. These results suggest that serum IDO activity can be used as a novel prognostic marker in PTB.

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Tryptophan 2,3-Dioxygenase Expression Identified in Murine Decidual Stromal Cells Is Not Essential for Feto-Maternal Tolerance

Frontiers in Immunology ◽

10.3389/fimmu.2020.601759 ◽

2020 ◽

Vol 11 ◽

Author(s):

Delia Hoffmann ◽

Tereza Dvorakova ◽

Florence Schramme ◽

Vincent Stroobant ◽

Benoit J. Van den Eynde

Keyword(s):

Stromal Cells ◽

Kynurenine Pathway ◽

Immune Rejection ◽

Cell Type ◽

Rate Limiting Step ◽

Limiting Step ◽

Tryptophan Catabolism ◽

Maternal Tolerance ◽

Rate Limiting

Indoleamine 2,3-dioxygenase 1 (IDO1) and tryptophan 2,3-dioxygenase (TDO) catalyze the rate-limiting step of tryptophan catabolism along the kynurenine pathway, which has important immuno suppressive properties, particularly in tumor cells and dendritic cells. The prominent expression of IDO1 in the placenta also suggested a role in preventing immune rejection of fetal tissues, and pharmacological inhibition of IDO1 induced abortion of allogeneic fetuses in mice. However, this was later challenged by the lack of rejection of allogeneic fetuses in IDO1-KO mice, suggesting that other mechanisms may compensate for IDO1 deficiency. Here we investigated whether TDO could contribute to feto-maternal tolerance and compensate for IDO1 deficiency in IDO1-KO mice. Expression of TDO mRNA was previously detected in placental tissues. We developed a new chimeric rabbit anti-TDO antibody to confirm TDO expression at the protein level and identify the positive cell type by immunohistochemistry in murine placenta. We observed massive TDO expression in decidual stromal cells, starting at day E3.5, peaking at day E6.5 then declining rapidly while remaining detectable until gestation end. IDO1 was also induced in decidual stromal cells, but only at a later stage of gestation when TDO expression declined. To determine whether TDO contributed to feto-maternal tolerance, we mated TDO-KO and double IDO1-TDO-KO females with allogeneic males. However, we did not observe reduced fertility. These results suggest that, despite its expression in decidual stromal cells, TDO is not a dominant mechanism of feto-maternal tolerance able to compensate for the absence of IDO1. Redundant additional mechanisms of immunosuppression likely take over in these KO mice. The massive expression of TDO during decidualization might suggest a role of TDO in angiogenesis or vessel tonicity, as previously described for IDO1.

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Functional and structural roles of the glutathione-binding residues in maize (Zea mays) glutathione S-transferase I

Biochemical Journal ◽

10.1042/bj3580101 ◽

2001 ◽

Vol 358 (1) ◽

pp. 101-110 ◽

Cited By ~ 28

Author(s):

Nikolaos E. LABROU ◽

Luciane V. MELLO ◽

Yannis D. CLONIS

Keyword(s):

Steady State ◽

Catalytic Mechanism ◽

Limited Proteolysis ◽

Glutathione S Transferase ◽

Rate Limiting Step ◽

Limiting Step ◽

Steady State Kinetics ◽

Helical Segment ◽

Α Helix ◽

Rate Limiting

The isoenzyme glutathione S-transferase (GST) I from maize (Zea mays) was cloned and expressed in Escherichia coli, and its catalytic mechanism was investigated by site-directed mutagenesis and dynamic studies. The results showed that the enzyme promotes proton dissociation from the GSH thiol and creates a thiolate anion with high nucleophilic reactivity by lowering the pKa of the thiol from 8.7 to 6.2. Steady-state kinetics fit well to a rapid equilibrium, random sequential Bi Bi mechanism, with intrasubunit modulation between the GSH binding site (G-site) and the electrophile binding site (H-site). The rate-limiting step of the reaction is viscosity-dependent, and thermodynamic data suggest that product release is rate-limiting. Five residues of GST I (Ser11, His40, Lys41, Gln53 and Ser67), which are located in the G-site, were individually replaced with alanine and their structural and functional roles in the 1-chloro-2,4-dinitrobenzene (CDNB) conjugation reaction were investigated. On the basis of steady-state kinetics, difference spectroscopy and limited proteolysis studies it is concluded that these residues: (1) contribute to the affinity of the G-site for GSH, as they are involved in side-chain interaction with GSH; (2) influence GSH thiol ionization, and thus its reactivity; (3) participate in kcat regulation by affecting the rate-limiting step of the reaction; and (4) in the cases of His40, Lys41 and Gln53 play an important role in the structural integrity of, and probably in the flexibility of, the highly mobile short 310-helical segment of α-helix 2 (residues 35–46), as shown by limited proteolysis experiments. These structural perturbations are probably transmitted to the H-site through changes in Phe35 conformation. This accounts for the modulation of Kcdnbm by His40, Lys41 and Gln53, and also for the intrasubunit communication between the G- and H-sites. Computer simulations using CONCOORD were applied to maize GST I monomer and dimer structures, each with bound lactoylglutathione, and the results were analysed by the essential dynamics technique. Differences in dynamics were found between the monomer and the dimer simulations showing the importance of using the whole structure in dynamic analysis. The results obtained confirm that the short 310-helical segment of α-helix 2 (residues 35–46) undergoes the most significant structural rearrangements. These rearrangements are discussed in terms of enzyme catalytic mechanism.

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Solid products and rate-limiting step in the thermal half decomposition of natural dolomite in a CO2(g) atmosphere

Thermochimica Acta ◽

10.1016/s0040-6031(02)00603-2 ◽

2003 ◽

Cited By ~ 1

Author(s):

D Beruto

Keyword(s):

Rate Limiting Step ◽

Limiting Step ◽

Rate Limiting

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Ornithine Decarboxylase Activity in Cultured Endothelial Cells Stimulated by Serum, Thrombin and Serotonin

Thrombosis and Haemostasis ◽

10.1055/s-0038-1646709 ◽

1978 ◽

Vol 39 (02) ◽

pp. 496-503 ◽

Cited By ~ 8

Author(s):

P A D’Amore ◽

H B Hechtman ◽

D Shepro

Keyword(s):

Endothelial Cells ◽

Ornithine Decarboxylase ◽

Decarboxylase Activity ◽

Aortic Endothelial Cells ◽

Ornithine Decarboxylase Activity ◽

Rate Limiting Step ◽

Bovine Aortic Endothelial Cells ◽

Limiting Step ◽

Rate Limiting ◽

Serum Serotonin

SummaryOrnithine decarboxylase (ODC) activity, the rate-limiting step in the synthesis of polyamines, can be demonstrated in cultured, bovine, aortic endothelial cells (EC). Serum, serotonin and thrombin produce a rise in ODC activity. The serotonin-induced ODC activity is significantly blocked by imipramine (10-5 M) or Lilly 11 0140 (10-6M). Preincubation of EC with these blockers together almost completely depresses the 5-HT-stimulated ODC activity. These observations suggest a manner by which platelets may maintain EC structural and metabolic soundness.

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