Escherichia coli K-12 substr. MG1655 Enzyme: D-alanine-D-alanine ligase A

Gene: ddlA Accession Numbers: EG10213 (EcoCyc), b0381, ECK0376

Regulation Summary Diagram: ?

Regulation summary diagram for ddlA

DdlA is one of two D-alanine-D-alanine ligases in E. coli [Zawadzke91]. D-alanine-D-alanine ligase, along with alanine racemase, makes up the D-alanine branch of peptidoglycan biosynthesis. The enzyme synthesizes D-alanyl-D-alanine from two molecules of D-alanine, which is then added to the growing cell wall precursor.

D-alanine-D-alanine ligase is an antibacterial drug target; it has long known to be the target of D-cycloserine [Neuhaus64].

Locations: cytosol

Map Position: [399,053 <- 400,147] (8.6 centisomes, 31°)
Length: 1095 bp / 364 aa

Molecular Weight of Polypeptide: 39.316 kD (from nucleotide sequence), 40.0 kD (experimental) [Zawadzke91 ]

pI: 5.27

Isozyme Sequence Similarity:
D-alanine-D-alanine ligase B: YES

Unification Links: ASAP:ABE-0001309 , CGSC:30966 , DIP:DIP-47939N , EchoBASE:EB0209 , EcoGene:EG10213 , EcoliWiki:b0381 , ModBase:P0A6J8 , OU-Microarray:b0381 , PortEco:ddlA , PR:PRO_000022419 , Pride:P0A6J8 , Protein Model Portal:P0A6J8 , RefSeq:NP_414915 , RegulonDB:EG10213 , SMR:P0A6J8 , String:511145.b0381 , UniProt:P0A6J8

Relationship Links: InterPro:IN-FAMILY:IPR000291 , InterPro:IN-FAMILY:IPR005905 , InterPro:IN-FAMILY:IPR011095 , InterPro:IN-FAMILY:IPR011127 , InterPro:IN-FAMILY:IPR011761 , InterPro:IN-FAMILY:IPR013815 , InterPro:IN-FAMILY:IPR013816 , InterPro:IN-FAMILY:IPR016185 , Panther:IN-FAMILY:PTHR23132 , Pfam:IN-FAMILY:PF01820 , Pfam:IN-FAMILY:PF07478 , Prosite:IN-FAMILY:PS00843 , Prosite:IN-FAMILY:PS00844 , Prosite:IN-FAMILY:PS50975

In Paralogous Gene Group: 115 (2 members)

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

GO Terms:

Biological Process: GO:0008360 - regulation of cell shape Inferred by computational analysis [UniProtGOA11a]
GO:0009252 - peptidoglycan biosynthetic process Inferred by computational analysis [UniProtGOA12, UniProtGOA11a, GOA06]
GO:0071555 - cell wall organization Inferred by computational analysis [UniProtGOA11a]
Molecular Function: GO:0008716 - D-alanine-D-alanine ligase activity Inferred from experiment Inferred by computational analysis [GOA06, GOA01a, GOA01, alBar92, Zawadzke91]
GO:0000166 - nucleotide binding Inferred by computational analysis [UniProtGOA11a]
GO:0000287 - magnesium ion binding Inferred by computational analysis [GOA06]
GO:0003824 - catalytic activity Inferred by computational analysis [GOA01]
GO:0005524 - ATP binding Inferred by computational analysis [UniProtGOA11a, GOA06, GOA01]
GO:0016874 - ligase activity Inferred by computational analysis [UniProtGOA11a]
GO:0030145 - manganese ion binding Inferred by computational analysis [GOA06]
GO:0046872 - metal ion binding Inferred by computational analysis [UniProtGOA11a, GOA01]
Cellular Component: GO:0005829 - cytosol Inferred from experiment Inferred by computational analysis [DiazMejia09, Ishihama08, LopezCampistrou05]
GO:0005737 - cytoplasm Inferred by computational analysis [UniProtGOA11, UniProtGOA11a, GOA06, GOA01]

MultiFun Terms: cell structure murein
metabolism biosynthesis of macromolecules (cellular constituents) murein (peptidoglycan)

Essentiality data for ddlA knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB enriched Yes 37 Aerobic 6.95   Yes [Gerdes03, Comment 1]
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 2]
M9 medium with 1% glycerol Yes 37 Aerobic 7.2 0.35 Yes [Joyce06, Comment 3]
MOPS medium with 0.4% glucose Yes 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 2]
Yes [Feist07, Comment 4]

Last-Curated ? 15-Aug-2012 by Keseler I , SRI International

Enzymatic reaction of: D-alanine-D-alanine ligase A

Synonyms: D-alanylalanine synthetase, alanylalanine synthetase, alanine:alanine ligase (ADP-forming), DD-ligase

EC Number:

2 D-alanine + ATP <=> D-alanyl-D-alanine + ADP + phosphate + H+

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction in which it was curated.

The reaction is physiologically favored in the direction shown.

Alternative Substrates for D-alanine: (R)-2-aminobutanoate [Zawadzke91 ] , glycine [Zawadzke91 ]

In Pathways: peptidoglycan biosynthesis I (meso-diaminopimelate containing) , UDP-N-acetylmuramoyl-pentapeptide biosynthesis I (meso-diaminopimelate containing)

Inhibition of the enzyme by a variety of phosphonate and phosphinate dipeptide analogs has been studied [Ellsworth96].

Inhibitors (Competitive): D-cycloserine (Kic = 8.9µM) [Zawadzke91]

Inhibitors (Noncompetitive): D-alanyl-D-alanine (Ki = 49µM) [Lugtenberg72, Zawadzke91]

Inhibitors (Unknown Mechanism): an aminoalkylphosphinate [Zawadzke91]

Kinetic Parameters:

Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)
[Shi95a, BRENDA14]
40000.0, 38.0
[Zawadzke91, BRENDA14]
3.0, 4.0, 110.0, 1100.0, 2000.0, 8000.0
11.83, 14.7, 26.7
3.94, 4.9, 8.9, 2.96, 3.68, 6.68, 0.11, 0.13, 0.24, 0.011, 0.013, 0.024, 0.0059, 0.0074, 0.013, 0.0015, 0.0018, 0.0033
[Park96a, BRENDA14]
[Healy98, BRENDA14]

T(opt): 37 °C [BRENDA14, Lugtenberg72]

Sequence Features

Protein sequence of D-alanine-D-alanine ligase A with features indicated

Feature Class Location Citations Comment
Conserved-Region 145 -> 348
UniProt: ATP-grasp;
Nucleotide-Phosphate-Binding-Region 175 -> 230
UniProt: ATP; Non-Experimental Qualifier: by similarity;
Metal-Binding-Site 302
UniProt: Magnesium or manganese 1; Non-Experimental Qualifier: by similarity;
Metal-Binding-Site 315
UniProt: Magnesium or manganese 1; Non-Experimental Qualifier: by similarity;
Metal-Binding-Site 317
UniProt: Magnesium or manganese 2; Non-Experimental Qualifier: by similarity;

Gene Local Context (not to scale): ?

Gene local context diagram

Transcription Unit:

Transcription-unit diagram


10/20/97 Gene b0381 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10213; confirmed by SwissProt match.


alBar92: al-Bar OA, O'Connor CD, Giles IG, Akhtar M (1992). "D-alanine: D-alanine ligase of Escherichia coli. Expression, purification and inhibitory studies on the cloned enzyme." Biochem J 282 ( Pt 3);747-52. PMID: 1554356

Baba06: Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, Datsenko KA, Tomita M, Wanner BL, Mori H (2006). "Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection." Mol Syst Biol 2;2006.0008. PMID: 16738554

BRENDA14: BRENDA team (2014). "Imported from BRENDA version existing on Aug 2014."

DiazMejia09: Diaz-Mejia JJ, Babu M, Emili A (2009). "Computational and experimental approaches to chart the Escherichia coli cell-envelope-associated proteome and interactome." FEMS Microbiol Rev 33(1);66-97. PMID: 19054114

Ellsworth96: Ellsworth BA, Tom NJ, Bartlett PA (1996). "Synthesis and evaluation of inhibitors of bacterial D-alanine:D-alanine ligases." Chem Biol 3(1);37-44. PMID: 8807826

Feist07: Feist AM, Henry CS, Reed JL, Krummenacker M, Joyce AR, Karp PD, Broadbelt LJ, Hatzimanikatis V, Palsson BO (2007). "A genome-scale metabolic reconstruction for Escherichia coli K-12 MG1655 that accounts for 1260 ORFs and thermodynamic information." Mol Syst Biol 3;121. PMID: 17593909

Gerdes03: Gerdes SY, Scholle MD, Campbell JW, Balazsi G, Ravasz E, Daugherty MD, Somera AL, Kyrpides NC, Anderson I, Gelfand MS, Bhattacharya A, Kapatral V, D'Souza M, Baev MV, Grechkin Y, Mseeh F, Fonstein MY, Overbeek R, Barabasi AL, Oltvai ZN, Osterman AL (2003). "Experimental determination and system level analysis of essential genes in Escherichia coli MG1655." J Bacteriol 185(19);5673-84. PMID: 13129938

GOA01: GOA, DDB, FB, MGI, ZFIN (2001). "Gene Ontology annotation through association of InterPro records with GO terms."

GOA01a: GOA, MGI (2001). "Gene Ontology annotation based on Enzyme Commission mapping." Genomics 74;121-128.

GOA06: GOA, SIB (2006). "Electronic Gene Ontology annotations created by transferring manual GO annotations between orthologous microbial proteins."

Healy98: Healy VL, Park IS, Walsh CT (1998). "Active-site mutants of the VanC2 D-alanyl-D-serine ligase, characteristic of one vancomycin-resistant bacterial phenotype, revert towards wild-type D-alanyl-D-alanine ligases." Chem Biol 5(4);197-207. PMID: 9545431

Ishihama08: Ishihama Y, Schmidt T, Rappsilber J, Mann M, Hartl FU, Kerner MJ, Frishman D (2008). "Protein abundance profiling of the Escherichia coli cytosol." BMC Genomics 9;102. PMID: 18304323

Joyce06: Joyce AR, Reed JL, White A, Edwards R, Osterman A, Baba T, Mori H, Lesely SA, Palsson BO, Agarwalla S (2006). "Experimental and computational assessment of conditionally essential genes in Escherichia coli." J Bacteriol 188(23);8259-71. PMID: 17012394

LopezCampistrou05: Lopez-Campistrous A, Semchuk P, Burke L, Palmer-Stone T, Brokx SJ, Broderick G, Bottorff D, Bolch S, Weiner JH, Ellison MJ (2005). "Localization, annotation, and comparison of the Escherichia coli K-12 proteome under two states of growth." Mol Cell Proteomics 4(8);1205-9. PMID: 15911532

Lugtenberg72: Lugtenberg EJ (1972). "Studies on Escherichia coli enzymes involved in the synthesis of uridine diphosphate-N-acetyl-muramyl-pentapeptide." J Bacteriol 110(1);26-34. PMID: 4552992

Neuhaus64: Neuhaus FC, Lynch JL (1964). "The enzymatic synthesis of D-alanyl-D-alanine. 3. On the inhibition of D-alanyl-D-alanine synthetase by the antibiotic D-cycloserine." Biochemistry 3;471-80. PMID: 14188160

Park96a: Park IS, Lin CH, Walsh CT (1996). "Gain of D-alanyl-D-lactate or D-lactyl-D-alanine synthetase activities in three active-site mutants of the Escherichia coli D-alanyl-D-alanine ligase B." Biochemistry 35(32);10464-71. PMID: 8756703

Shi95a: Shi Y, Walsh CT (1995). "Active site mapping of Escherichia coli D-Ala-D-Ala ligase by structure-based mutagenesis." Biochemistry 34(9);2768-76. PMID: 7893688

UniProt09: UniProt Consortium (2009). "UniProt version 15.8 released on 2009-10-01 00:00:00." Database.

UniProt10: UniProt Consortium (2010). "UniProt version 2010-07 released on 2010-06-15 00:00:00." Database.

UniProtGOA11: UniProt-GOA (2011). "Gene Ontology annotation based on the manual assignment of UniProtKB Subcellular Location terms in UniProtKB/Swiss-Prot entries."

UniProtGOA11a: UniProt-GOA (2011). "Gene Ontology annotation based on manual assignment of UniProtKB keywords in UniProtKB/Swiss-Prot entries."

UniProtGOA12: UniProt-GOA (2012). "Gene Ontology annotation based on UniPathway vocabulary mapping."

Zawadzke91: Zawadzke LE, Bugg TD, Walsh CT (1991). "Existence of two D-alanine:D-alanine ligases in Escherichia coli: cloning and sequencing of the ddlA gene and purification and characterization of the DdlA and DdlB enzymes." Biochemistry 1991;30(6);1673-82. PMID: 1993184

Other References Related to Gene Regulation

Oberto10: Oberto J (2010). "FITBAR: a web tool for the robust prediction of prokaryotic regulons." BMC Bioinformatics 11;554. PMID: 21070640

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Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
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