Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli BL21(DE3) cells | Chromobacterium violaceum |
Protein Variants | Comment | Organism |
---|---|---|
L101A | the mutant shows 26% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101C | the mutant shows 47% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101D | the mutant shows 5% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101E | the mutant shows 9% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101F | the mutant shows 133% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101G | the mutant shows 20% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101H | the mutant shows 16% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101I | the mutant shows 51% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101K | the mutant shows 29% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101M | the mutant shows 102% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101N | the mutant shows 15% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101P | the mutant shows 9% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101Q | the mutant shows 30% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101R | the mutant shows 29% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101S | the mutant shows 28% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101T | the mutant shows 26% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101V | the mutant shows 26% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101W | the mutant shows 55% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101Y | the mutant shows 153% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
L101Y/W180F | the double mutant displays higher L-tryptophan hydroxylation activity than the wild type enzyme with a 5.2fold increase in kcat | Chromobacterium violaceum |
W180A | the mutant shows 66% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180C | the mutant shows 119% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180D | the mutant shows 3% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180E | the mutant shows 6% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180F | the mutant shows 204% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180G | the mutant shows 8% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180H | the mutant shows 73% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180I | the mutant shows 113% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180K | the mutant shows 4% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180L | the mutant shows 174% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180M | the mutant shows 166% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180N | the mutant shows 49% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180P | the mutant shows 15% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180Q | the mutant shows 17% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180R | the mutant shows 85% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180S | the mutant shows 46% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180T | the mutant shows 44% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180V | the mutant shows 155% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
W180Y | the mutant shows 115% relative L-tryptophan hydroxylation activity compared to the wild type enzyme | Chromobacterium violaceum |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.033 | - |
L-phenylalanine | mutant enzyme W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.035 | - |
5,6,7,8-tetrahydrobiopterin | mutant enzyme W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.086 | - |
5,6,7,8-tetrahydrobiopterin | mutant enzyme L101Y/W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.099 | - |
5,6,7,8-tetrahydrobiopterin | mutant enzyme L101Y, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.111 | - |
L-phenylalanine | wild type enzyme, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.14 | - |
5,6,7,8-tetrahydrobiopterin | wild type enzyme, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.253 | - |
L-phenylalanine | mutant enzyme L101Y/W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.478 | - |
L-phenylalanine | mutant enzyme L101Y, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
1 | - |
L-tryptophan | mutant enzyme W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
3.44 | - |
L-tryptophan | wild type enzyme, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
3.54 | - |
L-tryptophan | mutant enzyme L101Y, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
4.06 | - |
L-tryptophan | mutant enzyme L101Y/W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Fe2+ | required for activity | Chromobacterium violaceum |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
34000 | - |
SDS-PAGE | Chromobacterium violaceum |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Chromobacterium violaceum | P30967 | - |
- |
Chromobacterium violaceum NBRC 12614 | P30967 | - |
- |
Purification (Comment) | Organism |
---|---|
Ni-TED 2000 column chromatography, gel filtration | Chromobacterium violaceum |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
0.307 | - |
mutant enzyme W180F, at 30°C, using L-phenylalanine as substrate | Chromobacterium violaceum |
0.342 | - |
mutant enzyme W180F, at 30°C, using 5,6,7,8-tetrahydrobiopterin as substrate | Chromobacterium violaceum |
0.347 | - |
wild type enzyme, at 30°C, using 5,6,7,8-tetrahydrobiopterin as substrate | Chromobacterium violaceum |
0.69 | - |
wild type enzyme, at 30°C, using L-tryptophan as substrate | Chromobacterium violaceum |
0.893 | - |
mutant enzyme W180F, at 30°C, using L-tryptophan as substrate | Chromobacterium violaceum |
1 | - |
mutant enzyme L101Y, at 30°C, using 5,6,7,8-tetrahydrobiopterin as substrate | Chromobacterium violaceum |
1 | - |
mutant enzyme L101Y/W180F, at 30°C, using L-phenylalanine as substrate | Chromobacterium violaceum |
1.05 | - |
mutant enzyme L101Y/W180F, at 30°C, using 5,6,7,8-tetrahydrobiopterin as substrate | Chromobacterium violaceum |
1.77 | - |
mutant enzyme L101Y, at 30°C, using L-tryptophan as substrate | Chromobacterium violaceum |
3.37 | - |
wild type enzyme, at 30°C, using L-phenylalanine as substrate | Chromobacterium violaceum |
3.62 | - |
mutant enzyme L101Y/W180F, at 30°C, using L-tryptophan as substrate | Chromobacterium violaceum |
10.16 | - |
mutant enzyme L101Y, at 30°C, using L-phenylalanine as substrate | Chromobacterium violaceum |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-phenylalanine + 5,6,7,8-tetrahydrobiopterin + O2 | - |
Chromobacterium violaceum | L-tyrosine + 4a-hydroxytetrahydrobiopterin | - |
? | |
L-phenylalanine + 5,6,7,8-tetrahydrobiopterin + O2 | - |
Chromobacterium violaceum NBRC 12614 | L-tyrosine + 4a-hydroxytetrahydrobiopterin | - |
? | |
L-tryptophan + 5,6,7,8-tetrahydrobiopterin + O2 | the activity for L-tryptophan is extremely low compared to L-phenylalanine activity levels | Chromobacterium violaceum | 5-hydroxy-L-tryptophan + 4a-hydroxytetrahydrobiopterin | - |
? | |
L-tryptophan + 5,6,7,8-tetrahydrobiopterin + O2 | the activity for L-tryptophan is extremely low compared to L-phenylalanine activity levels | Chromobacterium violaceum NBRC 12614 | 5-hydroxy-L-tryptophan + 4a-hydroxytetrahydrobiopterin | - |
? |
Synonyms | Comment | Organism |
---|---|---|
L-phenylalanine 4-hydroxylase | - |
Chromobacterium violaceum |
PAH | - |
Chromobacterium violaceum |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.18 | - |
L-phenylalanine | mutant enzyme W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.2 | - |
5,6,7,8-tetrahydrobiopterin | mutant enzyme W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.2 | - |
5,6,7,8-tetrahydrobiopterin | wild type enzyme, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.4 | - |
L-tryptophan | wild type enzyme, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.51 | - |
L-tryptophan | mutant enzyme W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.57 | - |
5,6,7,8-tetrahydrobiopterin | mutant enzyme L101Y, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.57 | - |
L-phenylalanine | mutant enzyme L101Y/W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
0.6 | - |
5,6,7,8-tetrahydrobiopterin | mutant enzyme L101Y/W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
1.02 | - |
L-tryptophan | mutant enzyme L101Y, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
1.93 | - |
L-phenylalanine | wild type enzyme, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
2.08 | - |
L-tryptophan | mutant enzyme L101Y/W180F, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum | |
5.83 | - |
L-phenylalanine | mutant enzyme L101Y, at 30°C, in 50 mM HEPES-NaOH buffer (pH 7.5) | Chromobacterium violaceum |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
5,6,7,8-tetrahydro-L-biopterin | - |
Chromobacterium violaceum |