Cloned (Comment) | Organism |
---|---|
gene bphD, DNA and amino acid sequence determination and analysis, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) | Dyella ginsengisoli |
Protein Variants | Comment | Organism |
---|---|---|
M148A | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148C | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148D | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148E | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148F | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148G | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148H | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148I | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148K | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148L | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148N | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148P | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148Q | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148R | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148S | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148T | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148V | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148W | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
M148Y | site-directed mutagenesis of the non-active-site residue | Dyella ginsengisoli |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten steady-state kinetics, stopped-flow measurements | Dyella ginsengisoli | |
0.00028 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148D | Dyella ginsengisoli | |
0.00044 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148S | Dyella ginsengisoli | |
0.00047 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148W | Dyella ginsengisoli | |
0.00058 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148R | Dyella ginsengisoli | |
0.00074 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148H | Dyella ginsengisoli | |
0.00075 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, wild-type enzyme | Dyella ginsengisoli | |
0.00078 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148I | Dyella ginsengisoli | |
0.00079 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148T | Dyella ginsengisoli | |
0.00097 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148C | Dyella ginsengisoli | |
0.0012 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148K | Dyella ginsengisoli | |
0.0012 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148V | Dyella ginsengisoli | |
0.0013 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148Q | Dyella ginsengisoli | |
0.0014 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148E | Dyella ginsengisoli | |
0.0014 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148L | Dyella ginsengisoli | |
0.0014 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148N | Dyella ginsengisoli | |
0.0019 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148Y | Dyella ginsengisoli | |
0.003 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148P | Dyella ginsengisoli | |
0.0042 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148F | Dyella ginsengisoli | |
0.0049 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148G | Dyella ginsengisoli | |
0.0051 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148A | Dyella ginsengisoli |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
32000 | - |
x * 32000, recombinant His-tagged enzyme, SDS-PAGE | Dyella ginsengisoli |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
2,6-dioxo-6-phenylhexa-3-enoate + H2O | Dyella ginsengisoli | - |
benzoate + 2-oxopent-4-enoate | - |
? | |
2,6-dioxo-6-phenylhexa-3-enoate + H2O | Dyella ginsengisoli LA-4 | - |
benzoate + 2-oxopent-4-enoate | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Dyella ginsengisoli | B5SU85 | gene bphD | - |
Dyella ginsengisoli LA-4 | B5SU85 | gene bphD | - |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography | Dyella ginsengisoli |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
2,6-dioxo-6-phenylhexa-3-enoate + H2O | - |
Dyella ginsengisoli | benzoate + 2-oxopent-4-enoate | - |
? | |
2,6-dioxo-6-phenylhexa-3-enoate + H2O | - |
Dyella ginsengisoli LA-4 | benzoate + 2-oxopent-4-enoate | - |
? | |
additional information | key role of non-active-site residue Met148 on the catalytic efficiency of meta-cleavage product hydrolase BphD | Dyella ginsengisoli | ? | - |
? | |
additional information | key role of non-active-site residue Met148 on the catalytic efficiency of meta-cleavage product hydrolase BphD | Dyella ginsengisoli LA-4 | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
? | x * 32000, recombinant His-tagged enzyme, SDS-PAGE | Dyella ginsengisoli |
Synonyms | Comment | Organism |
---|---|---|
BphD | - |
Dyella ginsengisoli |
MCP hydrolase | - |
Dyella ginsengisoli |
meta-cleavage product hydrolase | - |
Dyella ginsengisoli |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
20 | 25 | assay at | Dyella ginsengisoli |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.13 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148P | Dyella ginsengisoli | |
0.86 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148R | Dyella ginsengisoli | |
1.5 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148S | Dyella ginsengisoli | |
1.5 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148Y | Dyella ginsengisoli | |
2.4 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148K | Dyella ginsengisoli | |
2.5 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148C | Dyella ginsengisoli | |
2.6 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148H | Dyella ginsengisoli | |
3.1 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148L | Dyella ginsengisoli | |
4.1 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148E | Dyella ginsengisoli | |
4.5 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148D | Dyella ginsengisoli | |
4.8 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148V | Dyella ginsengisoli | |
5.7 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148W | Dyella ginsengisoli | |
6.5 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148T | Dyella ginsengisoli | |
6.7 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148N | Dyella ginsengisoli | |
8 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148F | Dyella ginsengisoli | |
8.4 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148Q | Dyella ginsengisoli | |
9.4 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148G | Dyella ginsengisoli | |
9.5 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148I | Dyella ginsengisoli | |
9.6 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, wild-type enzyme | Dyella ginsengisoli | |
9.9 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148A | Dyella ginsengisoli |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7 | - |
assay at | Dyella ginsengisoli |
General Information | Comment | Organism |
---|---|---|
additional information | structure analysis of wild-type and mutant enzymes, and molecular dynamics simulations of wild-type enzyme and mutants M148R and M148W. The catalytic triad residues of enzyme BphD are Ser112, Asp237, and His265. The most stable salt bridge between subunits of BphD wild-type and BphD mutant M148P is Glu434-Arg719, which has 58% and 85% occupied percentages in each trajectory, respectively | Dyella ginsengisoli |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
440 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148P | Dyella ginsengisoli | |
780 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148Y | Dyella ginsengisoli | |
1490 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148R | Dyella ginsengisoli | |
1900 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148F | Dyella ginsengisoli | |
1920 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148G | Dyella ginsengisoli | |
1940 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148A | Dyella ginsengisoli | |
2000 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148K | Dyella ginsengisoli | |
2210 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148L | Dyella ginsengisoli | |
2610 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148C | Dyella ginsengisoli | |
2880 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148E | Dyella ginsengisoli | |
3500 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148S | Dyella ginsengisoli | |
3590 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148H | Dyella ginsengisoli | |
4000 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148V | Dyella ginsengisoli | |
4790 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148N | Dyella ginsengisoli | |
6460 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148Q | Dyella ginsengisoli | |
8240 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148T | Dyella ginsengisoli | |
12100 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148W | Dyella ginsengisoli | |
12200 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148I | Dyella ginsengisoli | |
12800 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, wild-type enzyme | Dyella ginsengisoli | |
16100 | - |
2,6-dioxo-6-phenylhexa-3-enoate | pH 7.0, 25°C, mutant M148D | Dyella ginsengisoli |