Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli BL21(DE3) cells | Escherichia coli |
Crystallization (Comment) | Organism |
---|---|
hanging drop vapour diffusion method, using 100 mM Tris (pH 8.5), 200 mM NaCl, and 24% PEG 4000 | Escherichia coli |
Protein Variants | Comment | Organism |
---|---|---|
Q227A | the mutation causes an increase of kcat for uridine and inosine | Escherichia coli |
Q227F | the mutation causes an increase of kcat for uridine and inosine | Escherichia coli |
Q227Y | the mutation has a strong, enhancing effect on the hydrolysis of inosine, and the catalytic efficiency for the purinic substrate is increased by a factor of 7.6 | Escherichia coli |
T223A | the mutation does not improve significantly the catalytic efficiency of YeiK toward inosine | Escherichia coli |
T223F | the mutation does not improve significantly the catalytic efficiency of YeiK toward inosine | Escherichia coli |
T223F/Q227Y | the mutant shows a 2fold increase in catalytic efficiency toward inosine | Escherichia coli |
T223Y | the mutation does not affect the specificity of the enzyme toward inosine or uridine | Escherichia coli |
T223Y/Q227Y | the mutant displays a catalytic efficiency toward inosine that is more than 50fold increased compared to that of wild type enzyme | Escherichia coli |
T227A | the mutation does not improve significantly the catalytic efficiency of YeiK toward inosine | Escherichia coli |
T227F | the mutation does not improve significantly the catalytic efficiency of YeiK toward inosine | Escherichia coli |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.12 | - |
uridine | wild type enzyme, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.31 | - |
uridine | mutant enzyme T223F, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.33 | - |
uridine | mutant enzyme T223A, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.57 | - |
uridine | mutant enzyme Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.77 | - |
uridine | mutant enzyme Q227A, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
1.06 | - |
uridine | mutant enzyme T223Y/Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
1.09 | - |
uridine | mutant enzyme T223F/Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
1.13 | - |
uridine | mutant enzyme T223Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
1.16 | - |
Inosine | mutant enzyme T223Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
1.19 | - |
uridine | mutant enzyme Q227F, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
1.77 | - |
Inosine | mutant enzyme Q227F, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
1.93 | - |
Inosine | mutant enzyme T223Y/Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
2.14 | - |
Inosine | mutant enzyme Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
2.34 | - |
Inosine | wild type enzyme, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
3.29 | - |
Inosine | mutant enzyme T223A, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
4.31 | - |
Inosine | mutant enzyme Q227A, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
4.42 | - |
Inosine | mutant enzyme T223F/Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
5.3 | - |
Inosine | mutant enzyme T223F, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Ca2+ | the substrate binds to the Ca2+-containing active site in the catalytic cavity at the C-terminal end of the core beta-sheet | Escherichia coli |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Escherichia coli | P33022 | - |
- |
Purification (Comment) | Organism |
---|---|
Ni-NTA column chromatography and MonoQ column chromatography | Escherichia coli |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
inosine + H2O | poor substrate | Escherichia coli | hypoxanthine + D-ribose | - |
? | |
uridine + H2O | - |
Escherichia coli | uracil + D-ribose | - |
? |
Synonyms | Comment | Organism |
---|---|---|
CU-NH | - |
Escherichia coli |
cytidine-uridine-preferring nucleoside hydrolase | - |
Escherichia coli |
RihB | - |
Escherichia coli |
YeiK | - |
Escherichia coli |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.035 | - |
Inosine | mutant enzyme T223Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.086 | - |
Inosine | wild type enzyme, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.109 | - |
Inosine | mutant enzyme T223F, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.125 | - |
Inosine | mutant enzyme Q227F, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.18 | - |
Inosine | mutant enzyme T223A, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.182 | - |
Inosine | mutant enzyme Q227A, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.382 | - |
Inosine | mutant enzyme T223F/Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
0.593 | - |
Inosine | mutant enzyme Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
3.62 | - |
Inosine | mutant enzyme T223Y/Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
5.4 | - |
uridine | wild type enzyme, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
15.1 | - |
uridine | mutant enzyme Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
18.8 | - |
uridine | mutant enzyme T223F, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
39.8 | - |
uridine | mutant enzyme T223A, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
44.3 | - |
uridine | mutant enzyme T223Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
46.9 | - |
uridine | mutant enzyme Q227A, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
52.5 | - |
uridine | mutant enzyme Q227F, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
59.1 | - |
uridine | mutant enzyme T223F/Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli | |
72.9 | - |
uridine | mutant enzyme T223Y/Q227Y, in 50 mM HEPES buffer (pH 7.3), at 37°C | Escherichia coli |