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Literature summary for 3.5.1.18 extracted from

  • Nocek, B.; Starus, A.; Makowska-Grzyska, M.; Gutierrez, B.; Sanchez, S.; Jedrzejczak, R.; Mack, J.C.; Olsen, K.W.; Joachimiak, A.; Holz, R.C.
    The dimerization domain in DapE enzymes is required for catalysis (2014), PLoS ONE, 9, e93593.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 3.5.1.18

Cloned(Commentary)

Cloned (Comment) Organism
gene dapE, recombinant expression of N-terminally His6-tagged enzyme, containing a TEV protease recognition site followed by the DapE catalytic domain, in an Escherichia coli strain BL21(DE3) derivative that harbors the pMAGIC plasmid encoding one rare Escherichia coli Arg tRNA (covering codons AGG/AGA) Vibrio cholerae
gene dapE, recombinant expression of N-terminally His6-tagged wild-type and mutant enzymes, containing a TEV protease recognition site followed by the DapE catalytic domain, in an Escherichia coli strain BL21(DE3) derivative that harbors the pMAGIC plasmid encoding one rare Escherichia coli Arg tRNA (covering codons AGG/AGA) Haemophilus influenzae

Crystallization (Commentary)

Crystallization (Comment) Organism
purified recombinant enzyme, apoform and Zn2+-bound enzyme, using 400 nl of a precipitant solution containing 20% v/v 1,4-butanediol, 0.1 M sodium acetate, pH 4.5, and 400 nl of 19 mg/ml protein in crystallization buffer, with or without 1 mM ZnCl2, within 14 days, X-ray diffraction structure determination and analysis at 1.65 A resolution Vibrio cholerae
purified recombinant wild-type and mutant G172D enzymes, apoform and Zn2+-bound enzyme, using 400 nl of a precipitant solution containing 0.2 M ammonium acetate, 0.1 M Bis-Tris, pH 5.5, 25% w/v PEG 3350, and 400 nl of 15 mg/ml of protein in crystallization buffer, with or without 1 mM ZnCl2, within 14 days, X-ray diffraction structure determination and analysis at 1.84 A resolution Haemophilus influenzae

Protein Variants

Protein Variants Comment Organism
additional information construction of dimerization domain deletion mutants, that all show no activity Haemophilus influenzae
additional information construction of dimerization domain deletion mutants, that all show no activity Vibrio cholerae
T325A site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Haemophilus influenzae
T325C site-directed mutagenesis, inactive mutant Haemophilus influenzae
T325S site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme Haemophilus influenzae

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.8
-
 N-succinyl-LL-2,6-diaminoheptanedioate pH 7.5, 25°C, recombinant wild-type enzyme Haemophilus influenzae
1.2
-
 N-succinyl-LL-2,6-diaminoheptanedioate pH 7.5, 25°C, recombinant enzyme Vibrio cholerae
2.1
-
 N-succinyl-LL-2,6-diaminoheptanedioate pH 7.5, 25°C, recombinant mutant T325A Haemophilus influenzae
3
-
 N-succinyl-LL-2,6-diaminoheptanedioate pH 7.5, 25°C, recombinant mutant T325S Haemophilus influenzae

Metals/Ions

Metals/Ions Comment Organism Structure
Zn2+ required, dinuclear Zn(II)-loaded enzyme, binding structure, overview Haemophilus influenzae
Zn2+ required, dinuclear Zn(II)-loaded enzyme, binding structure, overview Vibrio cholerae

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
23200
-
 catalytic domain, dynamic light scattering Vibrio cholerae
28400
-
 catalytic domain, gel filtration Haemophilus influenzae
28700
-
 catalytic domain, dynamic light scattering Haemophilus influenzae
28700
-
 catalytic domain, gel filtration Vibrio cholerae
82600
-
 wild-type enzyme, gel filtration Haemophilus influenzae
83200
-
 wild-type enzyme, dynamic light scattering Haemophilus influenzae

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
N-succinyl-LL-2,6-diaminoheptanedioate + H2O Haemophilus influenzae
-
 succinate + LL-2,6-diaminoheptanedioate
-
 ?
N-succinyl-LL-2,6-diaminoheptanedioate + H2O Vibrio cholerae
-
 succinate + LL-2,6-diaminoheptanedioate
-
 ?

Organism

Organism UniProt Comment Textmining
Haemophilus influenzae
-
 gene dapE
-
Vibrio cholerae Q9KQ52 gene dapE
-

Purification (Commentary)

Purification (Comment) Organism
recombinant N-terminally His6-tagged enzyme from Escherichia coli strain BL21(DE3)/pMAGIC by nickel affinity chromatography and gel filtration Vibrio cholerae
recombinant N-terminally His6-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3)/pMAGIC by nickel affinity chromatography and gel filtration Haemophilus influenzae

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
N-succinyl-LL-2,6-diaminoheptanedioate + H2O
-
 Haemophilus influenzae succinate + LL-2,6-diaminoheptanedioate
-
 ?
N-succinyl-LL-2,6-diaminoheptanedioate + H2O
-
 Vibrio cholerae succinate + LL-2,6-diaminoheptanedioate
-
 ?

Subunits

Subunits Comment Organism
dimer the enzyme is composed of catalytic and dimerization domains, the dimerization domain in DapE enzymes is required for catalysis Haemophilus influenzae
dimer the enzyme is composed of catalytic and dimerization domains, the dimerization domain in DapE enzymes is required for catalysis Vibrio cholerae

Synonyms

Synonyms Comment Organism
DapE
-
 Haemophilus influenzae
DapE
-
 Vibrio cholerae
N-succinyl-L,L-diaminopimelic acid desuccinylase
-
 Haemophilus influenzae
N-succinyl-L,L-diaminopimelic acid desuccinylase
-
 Vibrio cholerae

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
 assay at Haemophilus influenzae
25
-
 assay at Vibrio cholerae

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.9
-
 N-succinyl-LL-2,6-diaminoheptanedioate pH 7.5, 25°C, recombinant mutant T325S Haemophilus influenzae
4
-
 N-succinyl-LL-2,6-diaminoheptanedioate pH 7.5, 25°C, recombinant mutant T325A Haemophilus influenzae
80
-
 N-succinyl-LL-2,6-diaminoheptanedioate pH 7.5, 25°C, recombinant enzyme Vibrio cholerae
114
-
 N-succinyl-LL-2,6-diaminoheptanedioate pH 7.5, 25°C, recombinant enzyme Haemophilus influenzae

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
 assay at Haemophilus influenzae
7.5
-
 assay at Vibrio cholerae

General Information

General Information Comment Organism
evolution the enzyme is composed of catalytic and dimerization domains, and belongs to the M20 peptidase family Haemophilus influenzae
evolution the enzyme is composed of catalytic and dimerization domains, and belongs to the M20 peptidase family Vibrio cholerae
additional information structural comparisons of wild-type and inactive monomeric DapE enzymes with other M20 peptidases, active site and zinc binding structure, molecular modeling and dynamics simulations, overview. The dimerization domain in DapE enzymes is required for catalysis. Removal of the dimerization domain increases the flexibility of a conserved active site loop that may provide critical interactions with the substrate Haemophilus influenzae
additional information structural comparisons of wild-type and inactive monomeric DapE enzymes with other M20 peptidases, active site and zinc binding structure, molecular modeling and dynamics simulations, overview. The dimerization domain in DapE enzymes is required for catalysis. Removal of the dimerization domain increases the flexibility of a conserved active site loop that may provide critical interactions with the substrate Vibrio cholerae