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Literature summary extracted from

  • Schnell, R.; Oehlmann, W.; Sandalova, T.; Braun, Y.; Huck, C.; Maringer, M.; Singh, M.; Schneider, G.
    Tetrahydrodipicolinate N-succinyltransferase and dihydrodipicolinate synthase from Pseudomonas aeruginosa: structure analysis and gene deletion (2012), PLoS ONE, 7, e31133.
    View publication on PubMedView publication on EuropePMC

Application

EC Number Application Comment Organism
2.3.1.117 additional information DapA is not an optimal target for drug development against Pseudomonas aeruginosa Pseudomonas aeruginosa
4.3.3.7 additional information the enzyme is not an optimal target for drug development against Pseudomonas aeruginosa Pseudomonas aeruginosa

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.3.1.117 gene dapD, phylogenetic tree, expression of His-tagged DapD in Escherichia coli strain BL21(DE3) Pseudomonas aeruginosa
4.3.3.7 gene dapA, expression of N-terminally His6-tagged enzyme in Escherichia coli strain BL21(DE3) Pseudomonas aeruginosa

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
2.3.1.117 purified recombinant His6-tagged DapD with bound L-2-aminopimelate and D-2-aminopimelate or in complex with CoA/succinate, hanging drop vapour diffusion method, mixing of 0.002 ml of 26 mg/ml protein in 25 mM Tris-HCl pH 8.0, and 150 mM NaCl, with or without 10-15 mM CoA, with 0.002 ml of reservoir solution containing 19-20% of PEG3350, 0.3-0.4 M succinate, pH 6.2, and equilibration against reservoir solution for 1-2 days, incubation of the enzyme with formyl-CoA leads to better crystals, soaking of apoenzyme crystals in solution containing L-2-aminopimelate and D-2-aminopimelate, the CoA-complex also contains a succinatemolecule bound next to the acceptor arm of the CoA in the active site cleft, X-ray diffraction structure determination and analysis at 1.8-2.95 A resolution, molecular replacement Pseudomonas aeruginosa
4.3.3.7 purified recombinant His6-tagged enzyme, hanging drop vapour diffusion method, mixing of 0.002 m of 12.5 mg/ml protein solution with 0.002 ml of reservoir solution containing 18% of PEG6000, 0.2 M MgCl2, and 0.1 M TRIS-HCl, pH 7.6, X-ray diffraction structure determination and analysis at 1.6 A resolution, molecular replacement Pseudomonas aeruginosa

Protein Variants

EC Number Protein Variants Comment Organism
4.3.3.7 additional information mutant construction by gene replacement of gene dapA (PA1010) via the sacB-based strategy Pseudomonas aeruginosa

Inhibitors

EC Number Inhibitors Comment Organism Structure
2.3.1.117 D-2-aminopimelate very weak competitive inhibition, L-2-aminopimelate and D-2-aminopimelate bind at the same site of the enzyme. Binding interaction analysis of the ligands in the enzyme active site suggests a misalignment of the amino group of D-2-aminopimelate for nucleophilic attack on the succinate moiety of the co-substrate succinyl-CoA as the structural basis of specificity and inhibition Pseudomonas aeruginosa

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.3.1.117 additional information
-
 additional information reaction kinetics for DapD, overview Pseudomonas aeruginosa
2.3.1.117 7
-
 (S)-2,3,4,5-tetrahydropyridine-2,6-dicarboxylate pH 7.5, 22°C Pseudomonas aeruginosa

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.3.1.117 Mg2+
-
 Pseudomonas aeruginosa

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
2.3.1.117 110000
-
 recombinant DapD, gel filtration Pseudomonas aeruginosa

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.3.1.117 succinyl-CoA + (S)-2,3,4,5-tetrahydropyridine-2,6-dicarboxylate + H2O Pseudomonas aeruginosa the enzyme is absolutely specific for the L-2-aminopimelate enantiomer CoA + N-succinyl-L-2-amino-6-oxoheptanedioate
-
 ?
4.3.3.7 (S)-aspartate-4-semialdehyde + pyruvate Pseudomonas aeruginosa
-
 (2S,4S)-4-hydroxy-2,3,4,5-tetrahydrodipicolinic acid + H2O
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.3.1.117 Pseudomonas aeruginosa
-
 gene dapD
-
4.3.3.7 Pseudomonas aeruginosa Q9I4W3 gene dapA or PA1010
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.3.1.117 recombinant His-tagged DapD from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, removal of te the N-terminal His6-tag by thrombin cleavage is not successful Pseudomonas aeruginosa
4.3.3.7 recombinant His6-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, the proteolytic cleavage by TEV protease does not remove the N-terminal His6-tag efficiently Pseudomonas aeruginosa

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.3.1.117 additional information no activity ith L-lysine, adipic acid, alpha-amino-adipic acid, L-epsilon-acetyl-lysine, L-glutamate, L-glutamine, L-norleucine, substrate specificity for DapD, overview. Binding of CoA to PaDapD does not induce any large conformational changes, ternary complex structure of DapD with bound CoA and succinate, overview Pseudomonas aeruginosa ?
-
 ?
2.3.1.117 succinyl-CoA + (S)-2,3,4,5-tetrahydropyridine-2,6-dicarboxylate + H2O the enzyme is absolutely specific for the L-2-aminopimelate enantiomer Pseudomonas aeruginosa CoA + N-succinyl-L-2-amino-6-oxoheptanedioate
-
 ?
2.3.1.117 succinyl-CoA + (S)-2,3,4,5-tetrahydropyridine-2,6-dicarboxylate + H2O the enzyme is absolutely specific for the L-2-aminopimelate enantiomer, L-2-aminopimelate and weak inhibitor D-2-aminopimelate bind at the same site of the enzyme. Binding interaction analysis of the ligands in the enzyme active site suggests a misalignment of the amino group of D-2-aminopimelate for nucleophilic attack on the succinate moiety of the co-substrate succinyl-CoA as the structural basis of specificity and inhibition Pseudomonas aeruginosa CoA + N-succinyl-L-2-amino-6-oxoheptanedioate
-
 ?
4.3.3.7 (S)-aspartate-4-semialdehyde + pyruvate
-
 Pseudomonas aeruginosa (2S,4S)-4-hydroxy-2,3,4,5-tetrahydrodipicolinic acid + H2O
-
 ?

Subunits

EC Number Subunits Comment Organism
2.3.1.117 trimer the subunit of PaDapD consists of three domains, the N-terminal globular domain, a central domain, and a C-terminal domain, overview Pseudomonas aeruginosa

Synonyms

EC Number Synonyms Comment Organism
2.3.1.117 DapD
-
 Pseudomonas aeruginosa
2.3.1.117 PA3666
-
 Pseudomonas aeruginosa
2.3.1.117 tetrahydrodipicolinate N-succinyltransferase
-
 Pseudomonas aeruginosa
4.3.3.7 DapA
-
 Pseudomonas aeruginosa
4.3.3.7 PA1010
-
 Pseudomonas aeruginosa

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.3.1.117 22
-
 assay at Pseudomonas aeruginosa
4.3.3.7 22
-
 assay at Pseudomonas aeruginosa

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.3.1.117 7.5
-
 assay at Pseudomonas aeruginosa
4.3.3.7 8
-
 assay at Pseudomonas aeruginosa

Cofactor

EC Number Cofactor Comment Organism Structure
2.3.1.117 succinyl-CoA
-
 Pseudomonas aeruginosa

IC50 Value

EC Number IC50 Value IC50 Value Maximum Comment Organism Inhibitor Structure
2.3.1.117 20
-
 pH 7.5, 22°C Pseudomonas aeruginosa D-2-aminopimelate

General Information

EC Number General Information Comment Organism
2.3.1.117 evolution the DAP biosynthesis pathway is present in most Gram-negative bacteria and mycobacteria Pseudomonas aeruginosa
2.3.1.117 metabolism tetrahydrodipicolinate N-succinyltransferase catalyses the transfer of the succinyl moiety of succinyl-CoA to the alpha-amino group of tetrahydrodipicolinate, the first committed step in the succinylase branch of the DAP biosynthesis pathway, diaminopimelic acid pathway of lysine biosynthesis, overview Pseudomonas aeruginosa
4.3.3.7 malfunction enzyme mutants with deleted dapA gene are viable and able to grow in a mouse lung infection model Pseudomonas aeruginosa
4.3.3.7 metabolism the enzyme catalyzes the first step in the diaminopimelic acid pathway of lysine biosynthesis Pseudomonas aeruginosa
4.3.3.7 additional information structure-based sequence alignments, based on the DapA crystal structure, reveal the presence of two homologues, PA0223 and PA4188, in Pseudomonas aeruginosa that can substitute for DapA in the PAO1DELTAdapA mutant. In vitro experiments using recombinant PA0223 protein do not detect any DapA activity Pseudomonas aeruginosa