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Literature summary for 3.7.1.8 extracted from

  • Zhou, H.; Qu, Y.; Kong, C.; Shen, E.; Wang, J.; Zhang, X.; Ma, Q.; Zhou, J.
    The key role of a non-active-site residue Met148 on the catalytic efficiency of meta-cleavage product hydrolase BphD (2013), Appl. Microbiol. Biotechnol., 97, 10399-10411.
    View publication on PubMed
Search for more literature for 3.7.1.8

Cloned(Commentary)

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

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 [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 [Da] Molecular Weight Maximum [Da] Comment Organism
32000
-
 x * 32000, recombinant His-tagged enzyme, SDS-PAGE Dyella ginsengisoli

Natural Substrates/ Products (Substrates)

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

Organism UniProt Comment Textmining
Dyella ginsengisoli B5SU85 gene bphD
-
Dyella ginsengisoli LA-4 B5SU85 gene bphD
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography Dyella ginsengisoli

Substrates and Products (Substrate)

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

Subunits Comment Organism
? x * 32000, recombinant His-tagged enzyme, SDS-PAGE Dyella ginsengisoli

Synonyms

Synonyms Comment Organism
BphD
-
 Dyella ginsengisoli
MCP hydrolase
-
 Dyella ginsengisoli
meta-cleavage product hydrolase
-
 Dyella ginsengisoli

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
20 25 assay at Dyella ginsengisoli

Turnover Number [1/s]

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

pH Optimum Minimum pH Optimum Maximum Comment Organism
7
-
 assay at Dyella ginsengisoli

General Information

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 [mM/s]

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