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Literature summary extracted from

  • Mimasu, S.; Yamagishi, H.; Kubo, S.; Kiyohara, M.; Matsuda, T.; Yahata, T.; Thomson, H.A.; Hupp, C.D.; Liu, J.; Okuda, T.; Kakefuda, K.
    Full-length in meso structure and mechanism of rat kynurenine 3-monooxygenase inhibition (2021), Commun. Biol., 4, 159 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.14.13.9 recombinant enzyme expression in HEK-293 cells Homo sapiens
1.14.13.9 sequence comparisons, recombinant expression of GST-tagged full-length KMO (1-478) from baculovirus vector containing a thrombin cleavage site inserted between GST and the N-terminus of KMO, and a TEV site followed by a FLAG tag at the C-terminus with the Bac-to-Bac Baculovirus system in Spodoptera frugiperda (Sf9) cells. Recombinant enzyme expression in HEK-293 cells Rattus norvegicus

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.14.13.9 purified full-length structure of KMO in its membrane-embedded form complexed with inhibitors 2-(benzyloxy)-5-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid or 4-chloro-2-([5-chloro-2-(5-methoxy-1,3-dihydro-2H-isoindol-2-yl)-1,3-thiazole-4-carbonyl](methyl)amino)-5-fluorobenzoic acid, sitting drop vapour diffusion method, mixing of enzyme in lipidic cubic phase formed by mixing a 9:1 monoolein:cholesterol molten lipid mixture and inhibitors, with reservoir solution containing 0.04 M Tris-Cl, pH 7.0, 0.06 M Bis-Tris, pH 6.5, 0.3-0.51 M lithium sulfate, and 34-45% PEG 400 for 2-(benzyloxy)-5-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid and containing 0.04-0.05 M sodium citrate, pH 6.5, or 0.04 M Bis-Tris pH 6.5, 0.05-0.06 M Tris-Cl, pH 7.0, 0.12-0.43 M lithium sulfate, and 34-46% PEG 400 for 4-chloro-2-([5-chloro-2-(5-methoxy-1,3-dihydro-2H-isoindol-2-yl)-1,3-thiazole-4-carbonyl](methyl)amino)-5-fluorobenzoic acid, in a 2:3 ratio at 17°C for 1-3 days, X-ray diffraction structure determination and analysis at 3.0 A resolution Rattus norvegicus
1.14.13.9 purified full-length structure of KMO in its membrane-embedded form, complexed with 2-(benzyloxy)-5-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid and 4-chloro-2-([5-chloro-2-(5-methoxy-1,3-dihydro-2H-isoindol-2-yl)-1,3-thiazole-4-carbonyl](methyl)amino)-5-fluorobenzoic acid at 3.0 A resolution Homo sapiens

Protein Variants

EC Number Protein Variants Comment Organism
1.14.13.9 D184A site-directed mutagenesis, the mutation weakens the beta-sheet dimer interface of the enzyme Rattus norvegicus
1.14.13.9 Q187A site-directed mutagenesis, the mutation weakens the beta-sheet dimer interface of the enzyme Rattus norvegicus
1.14.13.9 R380A site-directed mutagenesis, the mutation has no effect on kynurenine hydroxylation, suggesting that residue R380 does not play a major role in substrate recognition Rattus norvegicus
1.14.13.9 Y185P site-directed mutagenesis, the mutation weakens the beta-sheet dimer interface of the enzyme Rattus norvegicus

Inhibitors

EC Number Inhibitors Comment Organism Structure
1.14.13.9 2-(benzyloxy)-5-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid
-
 Homo sapiens
1.14.13.9 2-(benzyloxy)-5-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid R380, a residue from the enzyme's C-terminal region, forms hydrogen bonds with the carboxylic acid moiety of the inhibitor, residues R85, Y99 and Y398 also form bonds to 2-(benzyloxy)-5-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid Rattus norvegicus
1.14.13.9 3-(4-methyl-5-phenyl-1H-pyrazol-1-yl)benzoic acid
-
 Homo sapiens
1.14.13.9 3-(4-methyl-5-phenyl-1H-pyrazol-1-yl)benzoic acid
-
 Rattus norvegicus
1.14.13.9 3-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid
-
 Homo sapiens
1.14.13.9 3-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid
-
 Rattus norvegicus
1.14.13.9 4-chloro-2-([5-chloro-2-(5-methoxy-1,3-dihydro-2H-isoindol-2-yl)-1,3-thiazole-4-carbonyl](methyl)amino)-5-fluorobenzoic acid
-
 Homo sapiens
1.14.13.9 4-chloro-2-([5-chloro-2-(5-methoxy-1,3-dihydro-2H-isoindol-2-yl)-1,3-thiazole-4-carbonyl](methyl)amino)-5-fluorobenzoic acid ligand-binding structure, overview Rattus norvegicus
1.14.13.9 6-[3-(4-chloro-3-fluorophenyl)pyridin-2-yl]-1-methylquinazoline-2,4(1H,3H)-dione
-
 Homo sapiens
1.14.13.9 6-[3-(4-chloro-3-fluorophenyl)pyridin-2-yl]-1-methylquinazoline-2,4(1H,3H)-dione
-
 Rattus norvegicus
1.14.13.9 additional information determinations of inhibition with the purified enzyme and a cell-based assay Homo sapiens
1.14.13.9 additional information determinations of inhibition with the purified enzyme and a cell-based assay, docking studies of KMO representative inhibitors, inhibition mechanism, overview Rattus norvegicus

Localization

EC Number Localization Comment Organism GeneOntology No. Textmining
1.14.13.9 mitochondrial outer membrane KMO has two transmembrane domains, KMO is actually a single-pass transmembrane protein, with the other transmembrane domain lying laterally along the membrane, where it forms part of the ligand-binding pocket. Membrane-bound structure, overview Homo sapiens 5741
-
1.14.13.9 mitochondrial outer membrane KMO has two transmembrane domains, KMO is actually a single-pass transmembrane protein, with the other transmembrane domain lying laterally along the membrane, where it forms part of the ligand-binding pocket. Membrane-bound structure, overview Rattus norvegicus 5741
-

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.14.13.9 L-kynurenine + NADPH + H+ + O2 Homo sapiens
-
 3-hydroxy-L-kynurenine + NADP+ + H2O
-
 ?
1.14.13.9 L-kynurenine + NADPH + H+ + O2 Rattus norvegicus
-
 3-hydroxy-L-kynurenine + NADP+ + H2O
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.14.13.9 Homo sapiens O15229
-
-
1.14.13.9 Rattus norvegicus O88867
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.14.13.9 recombinant FLAG/GST-tagged full-length KMO (1-478) from Spodoptera frugiperda (Sf9) cells by glutathione affinity chromatography, ultrafiltration, and desalting gel filtration, followed by tag cleavage through thrombin, gel filtration, anti-FLAG affinity imuno-chromatography, and again ultrafiltration and gel filtration Rattus norvegicus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.14.13.9 L-kynurenine + NADPH + H+ + O2
-
 Homo sapiens 3-hydroxy-L-kynurenine + NADP+ + H2O
-
 ?
1.14.13.9 L-kynurenine + NADPH + H+ + O2
-
 Rattus norvegicus 3-hydroxy-L-kynurenine + NADP+ + H2O
-
 ?

Subunits

EC Number Subunits Comment Organism
1.14.13.9 homodimer ligand docking and human KMO model construction, overview Homo sapiens
1.14.13.9 homodimer the rat KMO monomer consists of three domains: alpha + beta flavin adenine dinucleotide (FAD)-binding domain I, FPMO enzyme-conserved domain II containing six-stranded beta-sheets, and C-terminal domain III, which is predicted to be comprising two transmembrane domains, although the exact topology remains ambiguous. The C-terminal region is comprising only one transmembrane domain, with the other predicted transmembrane helix lying laterally along the membrane, this helix displays hydrophobic residues on the outer side Rattus norvegicus

Synonyms

EC Number Synonyms Comment Organism
1.14.13.9 Hs-KMO
-
 Homo sapiens
1.14.13.9 KMO
-
 Homo sapiens
1.14.13.9 KMO
-
 Rattus norvegicus
1.14.13.9 Rat-KMO
-
 Rattus norvegicus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.14.13.9 37
-
 assay at Homo sapiens
1.14.13.9 37
-
 assay at Rattus norvegicus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.14.13.9 7.9
-
 assay at Homo sapiens
1.14.13.9 7.9
-
 assay at Rattus norvegicus

Cofactor

EC Number Cofactor Comment Organism Structure
1.14.13.9 FAD
-
 Homo sapiens
1.14.13.9 FAD
-
 Rattus norvegicus
1.14.13.9 NADPH
-
 Homo sapiens
1.14.13.9 NADPH
-
 Rattus norvegicus

IC50 Value

EC Number IC50 Value IC50 Value Maximum Comment Organism Inhibitor Structure
1.14.13.9 0.00000071
-
 pH 7.9, 37°C Homo sapiens 3-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid
1.14.13.9 0.0000017
-
 pH 7.9, 37°C Rattus norvegicus 4-chloro-2-([5-chloro-2-(5-methoxy-1,3-dihydro-2H-isoindol-2-yl)-1,3-thiazole-4-carbonyl](methyl)amino)-5-fluorobenzoic acid
1.14.13.9 0.0000038
-
 pH 7.9, 37°C Homo sapiens 2-(benzyloxy)-5-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid
1.14.13.9 0.0000068
-
 pH 7.9, 37°C Homo sapiens 6-[3-(4-chloro-3-fluorophenyl)pyridin-2-yl]-1-methylquinazoline-2,4(1H,3H)-dione
1.14.13.9 0.000009
-
 pH 7.9, 37°C Rattus norvegicus 2-(benzyloxy)-5-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid
1.14.13.9 0.000014
-
 pH 7.9, 37°C Homo sapiens 4-chloro-2-([5-chloro-2-(5-methoxy-1,3-dihydro-2H-isoindol-2-yl)-1,3-thiazole-4-carbonyl](methyl)amino)-5-fluorobenzoic acid
1.14.13.9 0.00013
-
 pH 7.9, 37°C Rattus norvegicus 6-[3-(4-chloro-3-fluorophenyl)pyridin-2-yl]-1-methylquinazoline-2,4(1H,3H)-dione
1.14.13.9 0.00015
-
 pH 7.9, 37°C Rattus norvegicus 3-[5-(4-chloro-3-fluorophenyl)-4-methyl-1H-pyrazol-1-yl]benzoic acid
1.14.13.9 0.00016
-
 pH 7.9, 37°C Homo sapiens 3-(4-methyl-5-phenyl-1H-pyrazol-1-yl)benzoic acid
1.14.13.9 0.00057
-
 pH 7.9, 37°C Rattus norvegicus 3-(4-methyl-5-phenyl-1H-pyrazol-1-yl)benzoic acid

General Information

EC Number General Information Comment Organism
1.14.13.9 malfunction mutations at the dimeric interface abolish the enzyme activity Homo sapiens
1.14.13.9 malfunction mutations at the dimeric interface abolish the enzyme activity Rattus norvegicus
1.14.13.9 additional information analysis of the extended ligand-binding pocket of in meso KMO and its binding mode Homo sapiens
1.14.13.9 additional information analysis of the extended ligand-binding pocket of in meso KMO and its binding mode Rattus norvegicus
1.14.13.9 physiological function kynurenine 3-monooxygenase (KMO) is a mitochondrial protein involved in the eukaryotic tryptophan catabolic pathway and is linked to various diseases Homo sapiens
1.14.13.9 physiological function kynurenine 3-monooxygenase (KMO) is a mitochondrial protein involved in the eukaryotic tryptophan catabolic pathway and is linked to various diseases Rattus norvegicus