Application | Comment | Organism |
---|---|---|
additional information | DapA is not an optimal target for drug development against Pseudomonas aeruginosa | Pseudomonas aeruginosa |
Cloned (Comment) | Organism |
---|---|
gene dapD, phylogenetic tree, expression of His-tagged DapD in Escherichia coli strain BL21(DE3) | Pseudomonas aeruginosa |
Crystallization (Comment) | Organism |
---|---|
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 |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
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] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | reaction kinetics for DapD, overview | Pseudomonas aeruginosa | |
7 | - |
(S)-2,3,4,5-tetrahydropyridine-2,6-dicarboxylate | pH 7.5, 22°C | Pseudomonas aeruginosa |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | - |
Pseudomonas aeruginosa |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
110000 | - |
recombinant DapD, gel filtration | Pseudomonas aeruginosa |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
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 | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Pseudomonas aeruginosa | - |
gene dapD | - |
Purification (Comment) | Organism |
---|---|
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 |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
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 | ? | - |
? | |
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 | - |
? | |
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 | - |
? |
Subunits | Comment | Organism |
---|---|---|
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 | Comment | Organism |
---|---|---|
DapD | - |
Pseudomonas aeruginosa |
PA3666 | - |
Pseudomonas aeruginosa |
tetrahydrodipicolinate N-succinyltransferase | - |
Pseudomonas aeruginosa |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
22 | - |
assay at | Pseudomonas aeruginosa |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.5 | - |
assay at | Pseudomonas aeruginosa |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
succinyl-CoA | - |
Pseudomonas aeruginosa |
IC50 Value | IC50 Value Maximum | Comment | Organism | Inhibitor | Structure |
---|---|---|---|---|---|
20 | - |
pH 7.5, 22°C | Pseudomonas aeruginosa | D-2-aminopimelate |
General Information | Comment | Organism |
---|---|---|
evolution | the DAP biosynthesis pathway is present in most Gram-negative bacteria and mycobacteria | Pseudomonas aeruginosa |
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 |