Cloned (Comment) | Organism |
---|---|
expresion in Escherichia coli and CHO cell | Homo sapiens |
Protein Variants | Comment | Organism |
---|---|---|
F31A/F34V/Q35H | methotrexate-resistant mutant, more than 4000fold decrease in ratio kcat/KM | Homo sapiens |
F31G/Q35N | methotrexate-resistant mutant, more than 50fold decrease in ratio kcat/KM | Homo sapiens |
F31P | methotrexate-resistant mutant, more than 30fold decrease in ratio kcat/KM | Homo sapiens |
F31P/Q35E | methotrexate-resistant mutant, more than 200fold decrease in ratio kcat/KM | Homo sapiens |
F31P/Q35H | methotrexate-resistant mutant, more than 150fold decrease in ratio kcat/KM | Homo sapiens |
F31R/F34A/Q35N | methotrexate-resistant mutant, more than 5000fold decrease in ratio kcat/KM | Homo sapiens |
F31R/F34T/Q35R | methotrexate-resistant mutant, more than 7000fold decrease in ratio kcat/KM | Homo sapiens |
F31R/F34T/Q35S | methotrexate-resistant mutant, more than 2000fold decrease in ratio kcat/KM | Homo sapiens |
F31R/Q35E | methotrexate-resistant mutant, more than 70fold decrease in ratio kcat/KM | Homo sapiens |
F31S/Q35E | methotrexate-resistant mutant, more than 450fold decrease in ratio kcat/KM | Homo sapiens |
additional information | characterization of ten methotrexate-resistant mutants with different amino acid combinations at residues F31, F34, and Q35. Higher resistance roughly correlates with a greater number of mutations, and an inverse correlation is observed between resistance and catalytic efficiency. Methotrexate-resistant dihydrofolate reductase can protect eucaryotic cells from methotrexate toxicity after transfection into dihydrofolate reductase knock-out cells | Homo sapiens |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
methotrexate | - |
Homo sapiens |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.000075 | - |
7,8-dihydrofolate | wild-type, pH 7.6, 23°C | Homo sapiens | |
0.00069 | - |
7,8-dihydrofolate | mutant F31R/Q35E, pH 7.6, 23°C | Homo sapiens | |
0.0015 | - |
7,8-dihydrofolate | mutant F31P, pH 7.6, 23°C | Homo sapiens | |
0.0017 | - |
7,8-dihydrofolate | mutant F31G/Q35N, pH 7.6, 23°C | Homo sapiens | |
0.0017 | - |
7,8-dihydrofolate | mutant F31R/F34T/Q35S, pH 7.6, 23°C | Homo sapiens | |
0.002 | - |
7,8-dihydrofolate | mutant F31R/F34T/Q35R, pH 7.6, 23°C | Homo sapiens | |
0.0021 | - |
7,8-dihydrofolate | mutant F31P/Q35H, pH 7.6, 23°C | Homo sapiens | |
0.0022 | - |
7,8-dihydrofolate | mutant F31R/F34A/Q35N, pH 7.6, 23°C | Homo sapiens | |
0.0027 | - |
7,8-dihydrofolate | mutant F31P/Q35E, pH 7.6, 23°C | Homo sapiens | |
0.0033 | - |
7,8-dihydrofolate | mutant F31S/Q35E, pH 7.6, 23°C | Homo sapiens | |
0.0043 | - |
7,8-dihydrofolate | mutant F31A/F34V/Q35H, pH 7.6, 23°C | Homo sapiens |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Homo sapiens | P00374 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
7,8-dihydrofolate + NADPH + H+ | - |
Homo sapiens | 5,6,7,8-tetrahydrofolate + NADP+ | - |
? |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.5 | - |
7,8-dihydrofolate | mutant F31R/F34A/Q35N, pH 7.6, 23°C | Homo sapiens | |
1 | - |
7,8-dihydrofolate | mutant F31R/F34T/Q35R, pH 7.6, 23°C | Homo sapiens | |
1 | - |
7,8-dihydrofolate | mutant F31R/F34T/Q35S, pH 7.6, 23°C | Homo sapiens | |
1.1 | - |
7,8-dihydrofolate | mutant F31A/F34V/Q35H, pH 7.6, 23°C | Homo sapiens | |
1.3 | - |
7,8-dihydrofolate | mutant F31R/Q35E, pH 7.6, 23°C | Homo sapiens | |
1.7 | - |
7,8-dihydrofolate | mutant F31P/Q35E, pH 7.6, 23°C | Homo sapiens | |
1.7 | - |
7,8-dihydrofolate | mutant F31P/Q35H, pH 7.6, 23°C | Homo sapiens | |
4.3 | - |
7,8-dihydrofolate | mutant F31G/Q35N, pH 7.6, 23°C | Homo sapiens | |
6.4 | - |
7,8-dihydrofolate | mutant F31P, pH 7.6, 23°C | Homo sapiens | |
9.4 | - |
7,8-dihydrofolate | mutant F31S/Q35E, pH 7.6, 23°C | Homo sapiens | |
10 | - |
7,8-dihydrofolate | wild-type, pH 7.6, 23°C | Homo sapiens |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
0.000000031 | - |
methotrexate | wild-type, pH 7.6, 23°C | Homo sapiens | |
0.0000017 | - |
methotrexate | mutant F31P, pH 7.6, 23°C | Homo sapiens | |
0.000009 | - |
methotrexate | mutant F31G/Q35N, pH 7.6, 23°C | Homo sapiens | |
0.00001 | - |
methotrexate | mutant F31P/Q35H, pH 7.6, 23°C | Homo sapiens | |
0.000012 | - |
methotrexate | mutant F31P/Q35E, pH 7.6, 23°C | Homo sapiens | |
0.000012 | - |
methotrexate | mutant F31R/F34A/Q35N, pH 7.6, 23°C | Homo sapiens | |
0.000021 | - |
methotrexate | mutant F31R/Q35E, pH 7.6, 23°C | Homo sapiens | |
0.00003 | - |
methotrexate | mutant F31S/Q35E, pH 7.6, 23°C | Homo sapiens | |
0.000059 | - |
methotrexate | mutant F31R/F34T/Q35S, pH 7.6, 23°C | Homo sapiens | |
0.000086 | - |
methotrexate | mutant F31R/F34T/Q35R, pH 7.6, 23°C | Homo sapiens | |
0.00018 | - |
methotrexate | mutant F31A/F34V/Q35H, pH 7.6, 23°C | Homo sapiens |
IC50 Value | IC50 Value Maximum | Comment | Organism | Inhibitor | Structure |
---|---|---|---|---|---|
0.000041 | - |
wild-type, pH 7.6, 23°C | Homo sapiens | methotrexate | |
0.00011 | - |
mutant F31P, pH 7.6, 23°C | Homo sapiens | methotrexate | |
0.00047 | - |
mutant F31P/Q35E, pH 7.6, 23°C | Homo sapiens | methotrexate | |
0.00047 | - |
mutant F31P/Q35H, pH 7.6, 23°C | Homo sapiens | methotrexate | |
0.00055 | - |
mutant F31R/F34A/Q35N, pH 7.6, 23°C | Homo sapiens | methotrexate | |
0.00057 | - |
mutant F31G/Q35N, pH 7.6, 23°C | Homo sapiens | methotrexate | |
0.00093 | - |
mutant F31S/Q35E, pH 7.6, 23°C | Homo sapiens | methotrexate | |
0.0031 | - |
mutant F31R/Q35E, pH 7.6, 23°C | Homo sapiens | methotrexate | |
0.0035 | - |
mutant F31R/F34T/Q35R, pH 7.6, 23°C | Homo sapiens | methotrexate | |
0.0044 | - |
mutant F31A/F34V/Q35H, pH 7.6, 23°C | Homo sapiens | methotrexate | |
0.0045 | - |
mutant F31R/F34T/Q35S, pH 7.6, 23°C | Homo sapiens | methotrexate |