Cloned (Comment) | Organism |
---|---|
gene DHFR, recombinant expression of His-tagged enzyme in Escherichia coli | Trypanosoma brucei brucei |
Crystallization (Comment) | Organism |
---|---|
purified recombinant His-tagged TbDHFR-TS, sitting drop vapour diffusion method, a few days at at 8°C or at room temperature, drops consist of 0.001 ml of protein solution and 0.001 ml of precipitant equilibrated against a 0.2 ml of reservoir solution. No crystal growth is observed over one year of incubation, preventing to obtain structural information on the enzyme | Trypanosoma brucei brucei |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
1-(3,4-dichlorophenyl)-6,6-dimethyl-1,6-dihydro-1,3,5-triazine-2,4-diamine | competitive inhibition | Trypanosoma brucei brucei | |
1-(3,4-dichlorophenyl)-6-(4-nitrophenyl)-1,6-dihydro-1,3,5-triazine-2,4-diamine | competitive inhibition | Trypanosoma brucei brucei | |
1-(4-chlorophenyl)-6-methyl-1,6-dihydro-1,3,5-triazine-2,4-diamine | competitive inhibition | Trypanosoma brucei brucei | |
1-(4-chlorophenyl)-6-propyl-1,6-dihydro-1,3,5-triazine-2,4-diamine | competitive inhibition | Trypanosoma brucei brucei | |
1-(4-methoxyphenyl)-6,6-dimethyl-1,6-dihydro-1,3,5-triazine-2,4-diamine | competitive inhibition | Trypanosoma brucei brucei | |
4-(4,6-diamino-2,2-dimethyl-1,3,5-triazin-1(2H)-yl)phenol | competitive inhibition | Trypanosoma brucei brucei | |
5-(4-chlorophenyl)-1,3,5-triazaspiro[5.5]undeca-1,3-diene-2,4-diamine | competitive inhibition | Trypanosoma brucei brucei | |
cycloguanil | CYC, the known inhibitor of Plasmodial and Trypanosomal DHFR-TS enzymes is also as an inhibitor of TbPTR1 | Trypanosoma brucei brucei | |
methotrexate | MTX | Trypanosoma brucei brucei | |
additional information | selectivity of the ihibitors versus enzymes PTR1 and DHFR, overview | Trypanosoma brucei brucei | |
N-[6-amino-5-(4-methoxyphenyl)-4,4-dimethyl-4,5-dihydrotriazin-2-yl]acetamide | competitive inhibition | Trypanosoma brucei brucei | |
pyrimethamine | PYR | Trypanosoma brucei brucei |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | kinetics by non-tight binding Michaelis-Menten model | Trypanosoma brucei brucei |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
7,8-dihydrobiopterin + NADPH + H+ | Trypanosoma brucei brucei | - |
5,6,7,8-tetrahydrobiopterin + NADP+ | - |
? | |
7,8-dihydrofolate + NADPH + H+ | Trypanosoma brucei brucei | - |
5,6,7,8-tetrahydrofolate + NADP+ | - |
? | |
biopterin + NADPH + H+ | Trypanosoma brucei brucei | reaction of EC 1.5.1.33 | 7,8-dihydrobiopterin + NADP+ | - |
? | |
folate + NADPH + H+ | Trypanosoma brucei brucei | - |
7,8-dihydrofolate + NADP+ | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Trypanosoma brucei brucei | Q27783 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged enzyme from Escherichia coli | Trypanosoma brucei brucei |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
7,8-dihydrobiopterin + NADPH + H+ | - |
Trypanosoma brucei brucei | 5,6,7,8-tetrahydrobiopterin + NADP+ | - |
? | |
7,8-dihydrofolate + NADPH + H+ | - |
Trypanosoma brucei brucei | 5,6,7,8-tetrahydrofolate + NADP+ | - |
? | |
biopterin + NADPH + H+ | reaction of EC 1.5.1.33 | Trypanosoma brucei brucei | 7,8-dihydrobiopterin + NADP+ | - |
? | |
folate + NADPH + H+ | - |
Trypanosoma brucei brucei | 7,8-dihydrofolate + NADP+ | - |
? |
Synonyms | Comment | Organism |
---|---|---|
bifunctional dihydrofolate reductase-thymidylate synthase | - |
Trypanosoma brucei brucei |
DHFR | - |
Trypanosoma brucei brucei |
DHFR-TS | - |
Trypanosoma brucei brucei |
dihydrofolate reductase | - |
Trypanosoma brucei brucei |
More | cf. EC 1.5.1.33 | Trypanosoma brucei brucei |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
22 | - |
assay at room temperature | Trypanosoma brucei brucei |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
10.5 | - |
assay at | Trypanosoma brucei brucei |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NADPH | - |
Trypanosoma brucei brucei |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | all the investigated molecules are assumed to act as full competitive inhibitor and analysed according to the non-tight binding Michaelis-Menten model | Trypanosoma brucei brucei |
IC50 Value | IC50 Value Maximum | Comment | Organism | Inhibitor | Structure |
---|---|---|---|---|---|
0.000024 | - |
pH 10.5, 22°C | Trypanosoma brucei brucei | pyrimethamine | |
0.00004 | - |
pH 10.5, 22°C | Trypanosoma brucei brucei | methotrexate | |
0.000103 | - |
pH 3.7, 22°C, recombinant enzyme | Trypanosoma brucei brucei | 1-(4-chlorophenyl)-6-methyl-1,6-dihydro-1,3,5-triazine-2,4-diamine | |
0.000256 | - |
pH 10.5, 22°C | Trypanosoma brucei brucei | cycloguanil | |
0.00726 | - |
pH 3.7, 22°C, recombinant enzyme | Trypanosoma brucei brucei | 1-(4-chlorophenyl)-6-propyl-1,6-dihydro-1,3,5-triazine-2,4-diamine | |
0.0176 | - |
pH 3.7, 22°C, recombinant enzyme | Trypanosoma brucei brucei | 5-(4-chlorophenyl)-1,3,5-triazaspiro[5.5]undeca-1,3-diene-2,4-diamine | |
0.0189 | - |
pH 3.7, 22°C, recombinant enzyme | Trypanosoma brucei brucei | N-[6-amino-5-(4-methoxyphenyl)-4,4-dimethyl-4,5-dihydrotriazin-2-yl]acetamide | |
0.0332 | - |
pH 3.7, 22°C, recombinant enzyme | Trypanosoma brucei brucei | 1-(4-methoxyphenyl)-6,6-dimethyl-1,6-dihydro-1,3,5-triazine-2,4-diamine | |
0.089 | - |
pH 3.7, 22°C, recombinant enzyme | Trypanosoma brucei brucei | 4-(4,6-diamino-2,2-dimethyl-1,3,5-triazin-1(2H)-yl)phenol |
General Information | Comment | Organism |
---|---|---|
evolution | PTR1 is a NADPH-dependent enzyme belonging to the short-chain dehydrogenase/reductase (SDR) family | Trypanosoma brucei brucei |
metabolism | traditional antifolates, such as methotrexate (MTX) inhibiting DHFR, are poorly effective towards trypanosome parasites because of the metabolic bypass provided by PTR1 also catalyzing folate reduction in addition to the conversion of biopterin to 7,8-dihydrobiopterin (DHB) and subsequently to 5,6,7,8-tetrahydrobiopterin (THB) | Trypanosoma brucei brucei |
physiological function | in addition to folate reduction, PTR1 (EC 1.5.1.33) also catalyzes the conversion of biopterin to 7,8-dihydrobiopterin (DHB) and subsequently to 5,6,7,8-tetrahydrobiopterin (THB). Under dihydrofolate reductase (DHFR) inhibition, PTR1 is upregulated providing reduced folates necessary for parasite survival | Trypanosoma brucei brucei |
physiological function | the bifunctional dihydrofolate reductase-thymidylate synthase, DHFR-TS, enzyme includes dthe ihydrofolate reductase, EC 1.5.1.3, and the thymidylate synthase, EC 2.1.1.45. Under dihydrofolate reductase (DHFR) inhibition, the PTR1 (EC 1.5.1.33) gene is upregulated, providing reduced folates necessary for parasite survival | Trypanosoma brucei brucei |