Application | Comment | Organism |
---|---|---|
drug development | the identification of a selective inhibitory allosteric binding site in the parasite enzyme and the fact that the human and trypanosome enzymes both display a strictly ordered bi-bi mechanism, but with the order of substrate binding reversed, makes the enzyme a good target for drug development | Trypanosoma brucei |
Cloned (Comment) | Organism |
---|---|
expression in Escherichia coli | Homo sapiens |
expression in Escherichia coli | Trypanosoma brucei brucei |
recombinant expression of His-tagged enzyme and of GST-tagged in Escherichia coli strain BL21(DE3) | Trypanosoma brucei |
Crystallization (Comment) | Organism |
---|---|
in complex with inhibitor 3-[2-(1,3-benzodioxol-5-yl)-2-oxoethyl]-4-bromo-3-hydroxy-5-methyl-1,3-dihydro-2H-indol-2-one. Only the (R)-enantiomer binds, and it is likely that the kinked shape of the molecule is crucial for its shape-complimentarity to the pocket. The benzo[1,3]dioxole moiety is deeply buried, making close contact with Ala397 and Gly232 at the bottom of the cleft. The indolin-2-one sits at the top of the cleft, with the unsubstituted edge exposed to solvent and the methyl and bromide substituents on making contact with Ala239, Met370, Lys371, and Ala367 | Trypanosoma brucei brucei |
purified recombinant detagged enzyme in complex with inhibitor 1, sitting-drop vapor diffusion method, mixing of 500 nl of 15 mg/ml protein solution wit 500 nl of precipitant solution containing 25% PEG 3350, 0.2 M (NH4)2SO4, 0.1 M Bis-Tris, pH 5.5, 22°C, X-ray diffraction structure determination and analysis at 1.75 A resolution | Trypanosoma brucei |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
3-[2-(1,3-benzodioxol-5-yl)-2-oxoethyl]-4-bromo-3-hydroxy-5-methyl-1,3-dihydro-2H-indol-2-one | competitive inhibitor with selectivity over the human counterpart, binds at an allosteric site absent in the human homologue that prevents the conformational rearrangement required to bind UTP | Trypanosoma brucei brucei | |
3-[2-(2,3-dihydro-1,4-benzodioxin-6-yl)-2-oxoethyl]-3-hydroxy-5-methyl-1,3-dihydro-2H-indol-2-one | - |
Trypanosoma brucei | |
3-[2-(2H-1,3-benzodioxol-5-yl)-2-oxoethyl]-3-hydroxy-6,7-dimethyl-1,3-dihydro-2H-indol-2-one | - |
Trypanosoma brucei | |
3-[2-(2H-1,3-benzodioxol-5-yl)-2-oxoethyl]-3-hydroxy-7-methyl-1,3-dihydro-2H-indol-2-one | - |
Trypanosoma brucei | |
3-[2-(2H-1,3-benzodioxol-5-yl)-2-oxoethyl]-4,6-dichloro-3-hydroxy-1,3-dihydro-2H-indol-2-one | - |
Trypanosoma brucei | |
3-[2H-(1,3-benzodioxol-5-yl)-2-oxoethyl]-4-bromo-3-hydroxy-5-methyl-1,3-dihydro-2H-indol-2-one | UTP-competitive inhibitor with selectivity over the human counterpart despite the high level of conservation of active site residues. The inhibitor binds at an allosteric site | Trypanosoma brucei | |
additional information | TbUAP inhibitor by high-throughput screening, and structure-activity relationships, overview. No inhibition by 3 | Trypanosoma brucei | |
N-(3,5-dimethoxyphenyl)-2,3-dihydro-4H-1,4-benzothiazine-4-carboxamide | - |
Trypanosoma brucei |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | the UAP enzyme kinetics display a strictly ordered bi.bi mechanism | Trypanosoma brucei |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | required | Trypanosoma brucei |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
UTP + N-acetyl-alpha-D-glucosamine 1-phosphate | Trypanosoma brucei | - |
diphosphate + UDP-N-acetyl-alpha-D-glucosamine | - |
r |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Homo sapiens | Q16222 | - |
- |
Trypanosoma brucei | - |
- |
- |
Trypanosoma brucei brucei | Q386Q8 | - |
- |
Trypanosoma brucei brucei 927 / 4 GUTat10.1 / TREU927 | Q386Q8 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, recombinant GST-tagged enzyme from Escherichia coli strain BL21(DE3) by glutathione affinity chromatography, tag cleavage, and gel filtration | Trypanosoma brucei |
Source Tissue | Comment | Organism | Textmining |
---|---|---|---|
bloodstream form | - |
Trypanosoma brucei | - |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
additional information | the human and trypanosome enzymes both display a strictly ordered bi-bi mechanism, but with the order of substrate binding reversed. Human UAP does not bind UTP alone, and although it does show significant binding to GlcNAc-1-P, it is not possible to calculate an affinity due to complex binding kinetics | Homo sapiens | ? | - |
? | |
additional information | the human and trypanosome enzymes both display a strictly ordered bi-bi mechanism, but with the order of substrate binding reversed. Trypanosoma brucei UAP binds UTP alone with a KD of 83.1 microM, but does not bind GlcNAc-1-P alone | Trypanosoma brucei brucei | ? | - |
? | |
additional information | the human and trypanosome enzymes both display a strictly ordered bi-bi mechanism, but with the order of substrate binding reversed. Trypanosoma brucei UAP binds UTP alone with a KD of 83.1 microM, but does not bind GlcNAc-1-P alone | Trypanosoma brucei brucei 927 / 4 GUTat10.1 / TREU927 | ? | - |
? | |
UTP + N-acetyl-alpha-D-glucosamine 1-phosphate | - |
Trypanosoma brucei | diphosphate + UDP-N-acetyl-alpha-D-glucosamine | - |
r |
Synonyms | Comment | Organism |
---|---|---|
Tb11.02.0120 | - |
Trypanosoma brucei brucei |
UAP | - |
Trypanosoma brucei |
uridine diphosphate N-acetylglucosamine pyrophosphorylase | - |
Trypanosoma brucei |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.5 | - |
assay at | Trypanosoma brucei |
IC50 Value | IC50 Value Maximum | Comment | Organism | Inhibitor | Structure |
---|---|---|---|---|---|
0.037 | - |
pH 7.5, temperature not specified in the publication | Trypanosoma brucei | 3-[2H-(1,3-benzodioxol-5-yl)-2-oxoethyl]-4-bromo-3-hydroxy-5-methyl-1,3-dihydro-2H-indol-2-one | |
0.037 | - |
pH not specified in the publication, temperature not specified in the publication | Trypanosoma brucei brucei | 3-[2-(1,3-benzodioxol-5-yl)-2-oxoethyl]-4-bromo-3-hydroxy-5-methyl-1,3-dihydro-2H-indol-2-one | |
0.049 | - |
pH 7.5, temperature not specified in the publication | Trypanosoma brucei | N-(3,5-dimethoxyphenyl)-2,3-dihydro-4H-1,4-benzothiazine-4-carboxamide |
General Information | Comment | Organism |
---|---|---|
physiological function | the enzyme catalyzes the final reaction in the biosynthesis of UDP-GlcNAc, an essential metabolite in many organisms including Trypanosoma brucei, | Trypanosoma brucei |