Literature summary for 6.3.2.4 extracted from

Huynh, K.H.; Hong, M.K.; Lee, C.; Tran, H.T.; Lee, S.H.; Ahn, Y.J.; Cha, S.S.; Kang, L.W.
The crystal structure of the D-alanine-D-alanine ligase from Acinetobacter baumannii suggests a flexible conformational change in the central domain before nucleotide binding (2015), J. Microbiol., 53, 776-782 .

Application

Application	Comment	Organism
drug development	the enzyme is a target for drug development. Acinetobacter baumannii is emerging as a multidrug-resistant nosocomial pathogen, that causes a number of diseases, including pneumonia, bacteremia, meningitis, and skin infections	Acinetobacter baumannii

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant enzyme, development of several different crystallization conditions for the AbDDL protein, e.g. sitting drop vapor diffusion method, 14°C, by mixing of 0.06 M MgCl2 and CaCl2, 0.1 M imidazole, 2-(N-morpholino)ethanesulfonic acid-HCl, pH 6.5, with 30% of the precipitant EDO-P8K containing 40% v/v ethylene glycol and 20% w/v PEG 8000, or by microbatch method with 0.2 M NaSCN and 20% w/v PEG 3350 as crystallization solution, X-ray diffraciton structure determination and analysis at 2.2 A resolution, molecular replacement wit the DDL crystal structure from Yestis pestis as a template, PDB ID 3v4z	Acinetobacter baumannii

Crystallization (Comment)

Organism

purified recombinant enzyme, development of several different crystallization conditions for the AbDDL protein, e.g. sitting drop vapor diffusion method, 14°C, by mixing of 0.06 M MgCl2 and CaCl2, 0.1 M imidazole, 2-(N-morpholino)ethanesulfonic acid-HCl, pH 6.5, with 30% of the precipitant EDO-P8K containing 40% v/v ethylene glycol and 20% w/v PEG 8000, or by microbatch method with 0.2 M NaSCN and 20% w/v PEG 3350 as crystallization solution, X-ray diffraciton structure determination and analysis at 2.2 A resolution, molecular replacement wit the DDL crystal structure from Yestis pestis as a template, PDB ID 3v4z

Acinetobacter baumannii

Inhibitors

Inhibitors	Comment	Organism
3-chloro-2,2-dimethyl-N-[4-(trifluoromethyl)phenyl]propanamide	binding structure, overview	Acinetobacter baumannii
D-cycloserine	-	Acinetobacter baumannii
additional information	proposed inhibitor binding site structure, overview	Acinetobacter baumannii
phosphorylated D-ala-D-alpha-hydroxybutyrate phosphonate	binding structure, overview	Acinetobacter baumannii

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
additional information	-	additional information	steady-state kinetics of the Michaelis-Menten equation with a single binding site	Acinetobacter baumannii
0.008	-	D-alanine	pH 7.8, 45°C, first D-Ala	Acinetobacter baumannii
1.7	-	ATP	pH 7.8, 45°C, first D-Ala	Acinetobacter baumannii
2.7	-	D-alanine	pH 7.8, 45°C, second D-Ala	Acinetobacter baumannii

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Mg2+	required	Acinetobacter baumannii

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
ATP + 2 D-alanine	Acinetobacter baumannii	-	ADP + phosphate + D-alanyl-D-alanine	-	?

Organism

Organism	UniProt	Comment	Textmining
Acinetobacter baumannii	-	-	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
ATP + 2 D-alanine	-	Acinetobacter baumannii	ADP + phosphate + D-alanyl-D-alanine	-	?

Subunits

Subunits	Comment	Organism
More	the asymmetric unit contains six protomers of AbDDL. Five protomers have a closed conformation in the central domain, while one protomer has an open conformation in the central domain. The central domain with an open conformation does not interact with crystallographic symmetry-related protomers and the conformational change of the central domain is not due to crystal packing. The central domain of AbDDL can have an ensemble of the open and closed conformations before the binding of substrate ATP. Active site structure determination and comparison, closed and open conformations of the central domain, overview	Acinetobacter baumannii

Synonyms

Synonyms	Comment	Organism
AbDDL	-	Acinetobacter baumannii
Ddl	-	Acinetobacter baumannii

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
45	-	assay at	Acinetobacter baumannii

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
30.08	-	D-alanine	pH 7.8, 45°C	Acinetobacter baumannii

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.8	-	assay at	Acinetobacter baumannii

Cofactor

Cofactor	Comment	Organism	Structure
ATP	the interdomain crevice between the central domain and the C-terminal domain contains the conserved nucleotide binding site, nucleotide binding site structure, overveiw	Acinetobacter baumannii

General Information

General Information	Comment	Organism
additional information	the crystal structure of the D-alanine-D-alanine ligase from Acinetobacter baumannii suggests a flexible conformational change in the central domain before nucleotide binding. The central domain of AbDDL can have an ensemble of the open and closed conformations before the binding of substrate ATP. The conformational change of the central domain is important for the catalytic activity. The conformational change does not result from crystal packing. The central domain is connected to the N-terminal and C-terminal domains through two parallel loops	Acinetobacter baumannii
physiological function	the D-alanine-D-alanine ligase (DDL) catalyzes the ATP-dependent synthesis of D-alanyl-D-alanine. D-Ala-D-ala is a key component of the bacterial cell wall and it maintains cell wall stability through cross-linking the peptide chain of peptidoglycan	Acinetobacter baumannii

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism	Structure
11.14	-	D-alanine	pH 7.8, 45°C, second D-Ala	Acinetobacter baumannii
3760.4	-	D-alanine	pH 7.8, 45°C, first D-Ala	Acinetobacter baumannii