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Information on EC 3.4.17.14 - Zinc D-Ala-D-Ala carboxypeptidase
for references in articles please use BRENDA:EC3.4.17.14
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EC Tree 3 Hydrolases
3.4 Acting on peptide bonds (peptidases)
3.4.17 Metallocarboxypeptidases
3.4.17.14 Zinc D-Ala-D-Ala carboxypeptidase
IUBMB Comments
A zinc enzyme. Catalyses carboxypeptidation but not transpeptidation reactions involved in bacterial cell wall metabolism. Weakly inhibited by β-lactams. In peptidase family M15. Distinct from EC 3.4.16.4, serine-type D-Ala-D-Ala carboxypeptidase.
The enzyme appears in viruses and cellular organisms
- 3.4.17.14
- peptidoglycan
- beta-lactams
-
exocellular
- benzylpenicillin
-
muropeptide
- actinomadura
- dd-peptidase
-
penicillin-sensitive
- dd-cpases
- murein
-
dd-endopeptidase
-
transpeptidation
- ld-carboxypeptidase
-
genapol
-
peptidoglycan-mimetic
-
bocillin
- homari
-
zn2+-containing
-
gaffkya
showSynonyms
dd-carboxypeptidase, pbp4a, d-alanyl-d-alanine carboxypeptidase, penicillin binding protein 4, dd-carboxypeptidase-transpeptidase, pbp4b, d-alanyl-d-alanine-cleaving carboxypeptidase, penicillin-binding protein 4a, more
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SYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
D-Alanyl-D-alanine hydrolase
-
-
-
-
D-Alanyl-D-alanine-cleaving carboxypeptidase
-
-
-
-
DD-Carboxypeptidase
DD-Carboxypeptidase-transpeptidase
-
-
-
-
G enzyme
-
-
-
-
Zn2+ G peptidase
-
-
-
-
DD-Carboxypeptidase
-
-
-
-
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REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cleavage of the bond: (Ac)2-L-lysyl-D-alanyl-/-D-alanine
-
-
-
-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
hydrolysis of peptide bond
hydrolysis of peptide bond
-
-
-
-
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PATHWAY SOURCE
PATHWAYS
MetaCyc
peptidoglycan biosynthesis V (beta-lactam resistance)
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CAS REGISTRY NUMBER
COMMENTARY
9077-67-2
-
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
LITERATURE
COMMENTARY
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
Nalpha,Nepsilon-diacetyl-Lys-D-Ala-D-Ala + H2O
Nalpha,Nepsilon-diacetyl-Lys-D-Ala + D-Ala
-
Substrates: -
Products: -
Products: -
?
peptidoglycan + H2O
?
-
Substrates: hydrolyzes peptidoglycan interpeptide bonds between a D-alanine and another D-amino acid residue in alpha-position to a free carboxylate
Products: -
Products: -
?
Acetyl2-L-Lys-D-Ala-D-Ala + H2O
Acetyl2-L-Lys-D-Ala + D-Ala
-
Substrates: -
Products: -
Products: -
?
Acetyl2-L-Lys-D-Ala-D-Ala + H2O
Acetyl2-L-Lys-D-Ala + D-Ala
-
Substrates: -
Products: -
Products: -
?
Acetyl2-L-Lys-D-Ala-D-Ala + H2O
Acetyl2-L-Lys-D-Ala + D-Ala
-
Substrates: -
Products: -
Products: -
?
Nalpha-Acetyl-L-Lys-D-Ala-D-Ala + H2O
Nalpha-Acetyl-L-Lys-D-Ala + D-Ala
-
Substrates: -
Products: -
Products: -
?
Nalpha-Acetyl-L-Lys-D-Ala-D-Ala + H2O
Nalpha-Acetyl-L-Lys-D-Ala + D-Ala
-
Substrates: -
Products: -
Products: -
?
additional information
?
-
-
Substrates: DAP contributes to resistance against NO required for the intracellular growth of the bacterium
Products: -
Products: -
?
additional information
?
-
-
Substrates: the DD-carboxypeptidase activity encoded by pbp4B is not essential for the cell growth of Escherichia coli
Products: -
Products: -
?
additional information
?
-
-
Substrates: PBP4B shows penicillin-binding capacity and a low DD-carboxypeptidase activity
Products: -
Products: -
?
additional information
?
-
-
Substrates: weak beta-lactamase activity hydrolyzing penicillin into penicilloate
Products: -
Products: -
?
additional information
?
-
-
Substrates: involved in bacterial cell wall metabolism
Products: -
Products: -
?
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NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
LITERATURE
COMMENTARY
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
additional information
?
-
-
Substrates: DAP contributes to resistance against NO required for the intracellular growth of the bacterium
Products: -
Products: -
?
additional information
?
-
-
Substrates: the DD-carboxypeptidase activity encoded by pbp4B is not essential for the cell growth of Escherichia coli
Products: -
Products: -
?
additional information
?
-
-
Substrates: involved in bacterial cell wall metabolism
Products: -
Products: -
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Zinc
Zn2+
Zinc
-
Zn2+ cofactor is required for both DD-carboxypeptidase activity and binding of benzyl-penicillin
Zinc
-
the Zn2+ ion is ligated by three histidine residues and located in a cleft in the C-terminal domain
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INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
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DISEASE
TITLE OF PUBLICATION
LINK TO PUBMED
Infections
Staphylococcus aureus cell wall structure and dynamics during host-pathogen interaction.
Osteomyelitis
Identification of Penicillin Binding Protein 4 (PBP4) as a critical factor for Staphylococcus aureus bone invasion during osteomyelitis in mice.
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.1
D-Ala-D-Ala
-
determined by isothermal titration calorimetry, at pH 8.0 and 37°C
0.14
D-Ala-D-Ala
-
determined by ninhydrin/Cd(II) assay, at pH 8.0 and 37°C
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
11.5
D-Ala-D-Ala
-
determined by isothermal titration calorimetry, at pH 8.0 and 37°C
43.8
D-Ala-D-Ala
-
determined by ninhydrin/Cd(II) assay, at pH 8.0 and 37°C
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
115
D-Ala-D-Ala
-
determined by isothermal titration calorimetry, at pH 8.0 and 37°C
312.9
D-Ala-D-Ala
-
determined by ninhydrin/Cd(II) assay, at pH 8.0 and 37°C
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Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0188
D-cysteine
-
determined by isothermal titration calorimetry, at pH 8.0 and 37°C
0.0232
D-cysteine
-
determined by ninhydrin/Cd(II) assay, at pH 8.0 and 37°C
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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pI VALUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain 544, ATCC 23448, a smooth virulent biovar 1 strain, gene dacF
-
-
brenda
the enzyme contains the mutation D261Y, relative to the SwissProt entry P24228; gene dacB
SwissProt
brenda
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SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
additional information
-
intracellular replication of Brucella abortus within murine macrophages and HeLa cells
brenda
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LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
the enzyme contains a 5' leader sequence targeting it to the periplasm
-
brenda
Highest Expressing Human Cell Lines
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UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). CBPM_STRAL
255
0
26190
Swiss-Prot
other Location (Reliability: 3)
DACB_ECOLI
Escherichia coli (strain K12)
477
0
51798
Swiss-Prot
other Location (Reliability: 2)
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MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
-
the enzyme consists of two globular domains, connected by a single link, the N-terminal domain has three alpha-helices, and the C-terminal domain has three alpha-helices and five beta-strands
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CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified recombinant enzyme, wild-type or selenomethionine-labeled, 10-15 mg/ml protein, hanging drop vapour diffusion method, from 0.1 M MES, pH 6.5, and 3-10% PEG 20000, microseeding, 20°C, cryoprotection by 25% v/v glycerol, X-ray diffraction structure determination and analysis at 1.55-1.70 A resolution, modeling
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
-
the enzyme mutant K23 of Brucella abortus fails to replicate in mouse macrophages and HeLa cells, and shows less virulence than the wild type in mice, Under nitric oxide (NO) stress, the growth of the DAP mutant in vitro decreases, and it also has less capability to reduce NO than the wild type, intracellular replication of the DAP mutant is partially restored by pretreatment of macrophages with the NO synthase inhibitor 1-phenyl-imidazole and the level of expression of the NO reductase gene norB in the DAP mutant is lower than that in the wild type
additional information
-
construction of gene pbp4B deletion mutants, the enzyme is not not essential for cell division, the strain lacking all known D-alanine carboxypeptidases is viable, overview
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PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant enzyme from strain BL21(DE3) by anion exchange and hydrophobic interaction chromatography, followed by gel filtration
recombinant His-tagged enzyme from strain BL21(DE3) membranes by solubilization with 1% Triton X-100 and nickel affinity chromatography
-
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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene dacB, expression of the enzyme lacking the 5' leader peptide in strain BL21(DE3)
gene pbp4B, DNA and amino acid sequence determination and analysis, the pbp4B gene is located at the zipA-dapA region, subcloning in strain DH5alpha, expression of N-terminally His-tagged enzyme in strain BL21(DE3) membranes
-
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REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Joris, B.; van Beeumen, J.; Casagrande, F.; Gerday, C.; Frere, J.M.; Ghuysen, J.M.
The complete amino acid sequence of the Zn2+-containing D-alanyl-D-alanine-cleaving carboxypeptidase of streptomyces albus G
Eur. J. Biochem.
130
53-69
1983
Streptomyces albus
brenda
Dideberg, O.; Charlier, P.; Dive, G.; Joris, B.; Frere, J.M.; Ghuysen, J.M.
Structure of a Zn2+-containing D-alanyl-D-alanine-cleaving carboxypeptidase at 2.5 A resolution
Nature
299
469-470
1982
Streptomyces albus
brenda
Dideberg, O.; Joris, B.; Frere, J.M.; Ghuysen, J.M.; Weber, G.; Robaye, R.; Delbrouck, J.M.; Roelandts, I.
The exocellular DD-carboxypeptidase of Streptomyces albus G: a metallo (Zn2+) enzyme
FEBS Lett.
117
215-218
1980
Streptomyces albus
brenda
Frere, J.M.; Moreno, R.; Ghuysen, J.M.; Perkins, H.R.; Dierickx, L.; Delcambe, L.
Molecular weight, amino acid composition and physicochemical properties of the exocellular DD-carboxypeptidase-transpeptidase of Streptomyces R39
Biochem. J.
143
233-240
1974
Streptomyces sp.
brenda
Kikuchi, H.; Kim, S.; Watanabe, K.; Watarai, M.
Brucella abortus d-alanyl-d-alanine carboxypeptidase contributes to its intracellular replication and resistance against nitric oxide
FEMS Microbiol. Lett.
259
120-125
2006
Brucella abortus
brenda
Ghuysen, J.
Zinc D-Ala-D-Ala carboxypeptidase (Streptomyces)
Handbook of Proteolytic Enzymes (Barrett,A. J. ;Rawlings,N. D. ,Woessner)
1
857-858
2004
Streptomyces albus G
-
brenda
Vega, D.; Ayala, J.A.
The DD-carboxypeptidase activity encoded by pbp4B is not essential for the cell growth of Escherichia coli
Arch. Microbiol.
185
23-27
2006
Escherichia coli
brenda
Kishida, H.; Unzai, S.; Roper, D.I.; Lloyd, A.; Park, S.Y.; Tame, J.R.
Crystal structure of penicillin binding protein 4 (dacB) from Escherichia coli, both in the native form and covalently linked to various antibiotics
Biochemistry
45
783-792
2006
Escherichia coli (P24228), Escherichia coli
brenda
Sauvage, E.; Duez, C.; Herman, R.; Kerff, F.; Petrella, S.; Anderson, J.W.; Adediran, S.A.; Pratt, R.F.; Frere, J.M.; Charlier, P.
Crystal structure of the Bacillus subtilis penicillin-binding protein 4a, and its complex with a peptidoglycan mimetic peptide
J. Mol. Biol.
371
528-539
2007
Bacillus subtilis
brenda
Lv, M.; Zhang, Y.J.; Zhou, F.; Ge, Y.; Zhao, M.H.; Liu, Y.; Yang, K.W.
Real-time monitoring of D-Ala-D-Ala dipeptidase activity of VanX in living bacteria by isothermal titration calorimetry
Anal. Biochem.
578
29-35
2019
Enterococcus faecalis
brenda
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