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Information on EC 2.3.2.B14 - L,D-transpeptidase and Organism(s) Mycobacterium tuberculosis and UniProt Accession O53223
for references in articles please use BRENDA:EC2.3.2.B14preliminary BRENDA-supplied EC number
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2.3.2.B14 L,D-transpeptidaseSpecify your search results
Word Map
- 2.3.2.B14
- tuberculosis
- mycobacterium
- carbapenems
-
penicillin-binding
-
d,d-transpeptidases
- enterococcus
- faecium
- ldtmt2
- transpeptidases
- imipenem
- ertapenem
- muropeptides
-
acylenzyme
- clavulanate
-
transpeptidation
- d-ala
-
lineage-determining
- doripenem
- d,d-carboxypeptidase
- analysis
The taxonomic range for the selected organisms is: Mycobacterium tuberculosis
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
Generates 3->3 cross-links in peptidoglycan, catalyzing the cleavage of the mDap(3)-D-Ala4 bond of a tetrapeptide donor stem and the formation of a bond between the carbonyl of mDap3 of the donor stem and the side chain of mDap3 of the acceptor stem.
Synonyms
l,d-transpeptidase, ldtmt1, ldtfs, l,d-transpeptidase 5, mt2594, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
SYSTEMATIC NAME
IUBMB Comments
peptidoglycan (3->3)-transpepdidase
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
2 GlcNAc-MurNAc-L-Ala-D-iGluNH2-meso-diaminopimelic acid-NH2-D-Ala
GlcNAc-MurNAc-L-Ala-D-iGluNH2-meso-diaminopimelic acid-NH2-D-Ala 3->3 crosslinked at the mesoDapNH2 position
-
-
-
-
?
nitrocefin + H2O
(2R)-2-[(R)-carboxy[2-(thiophen-2-yl)acetamido]methyl]-5-[(E)-2-(2,4-dinitrophenyl)ethenyl]-3,6-dihydro-2H-1,3-thiazine-4-carboxylic acid
-
-
-
?
additional information
?
-
the enzyme also shows low beta-lactamase activity with nitrocefin (i.e. 3-(2,4-dinitrostyryl)-(6R,7R)-7-(2-thienylacetamido)-ceph-3-em-4-carboxylic acid) with Km of 0.016 mM and kcat of 0.01 s-1 (in 20 mM HEPES (pH 7.5) and 300 mM NaCl, temperature not specified in the publication)
-
-
?
additional information
?
-
-
the enzyme also shows low beta-lactamase activity with nitrocefin (i.e. 3-(2,4-dinitrostyryl)-(6R,7R)-7-(2-thienylacetamido)-ceph-3-em-4-carboxylic acid) with Km of 0.016 mM and kcat of 0.01 s-1 (in 20 mM HEPES (pH 7.5) and 300 mM NaCl, temperature not specified in the publication)
-
-
?
additional information
?
-
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gene MT2594 (Rv2518c) encodes an L,Dtranspeptidase for synthesis of non-classical 3->3 cross-linkages
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-
?
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
biapenem
investigation of the binding interactions using a two-layered ONIOM model
tebipenem
investigation of the binding interactions using a two-layered ONIOM model
(4R,5S,6S)-3-[[(3S,5R)-5-(aminomethyl)oxolan-3-yl]sulfanyl]-6-[(1R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid
compound identified by virtual screening, interacts with residues Arg242 and Gly304
(4R,6S)-3-[[(3R,5R)-5-(aminomethyl)oxolan-3-yl]sulfanyl]-6-[(1R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid
2-[(2S,3S)-2-(2H-1,3-benzodioxol-5-yl)-1-(3-fluoro-4-methylphenyl)-4-oxoazetidin-3-yl]-5-nitro-1H-isoindole-1,3(2H)-dione
inhibitor identified by virtual screening, demonstrates close hydrogen bond interaction between the ligand and two active site residues Asn303 and Cys305, shows the highest binding free energy observed
2-[(2S,3S)-2-(4-fluorophenyl)-1-(3-methylphenyl)-4-oxoazetidin-3-yl]-5-nitro-1H-isoindole-1,3(2H)-dione
6-aminopenicillanic acid
high degree of Cys354 modification, occurrence of a possible coupling reaction
cefapirin
-
formation of a covalent comple that undergoes non-hydrolytic fragmentation, resulting in an adduct in which the C3' leaving group is lost
faropenem daloxate
high degree of Cys354 modification, fast degradation following the initial acylation event
Oxacillin
-
formation of a covalent comple that undergoes non-hydrolytic fragmentation. Cleavage of the C5-C6 bond of the penicillin-derived LdtB acyl-enzyme complex yields LdtB modified by an acetyl group bearing the penicillin C6 side chain
panipenem
panipenem is not degraded after binding. The presence of the 1-beta-methyl group in carbapenems is related to the ligand affinity of LdtB and the presence of the Y308 and Y318 residues in LdtB stabilizes the conformation of the LdtB-carbepenem adduct
sulopenem
-
formation of a covalent comple that undergoes non-hydrolytic fragmentation, yielding LdtB acylated with a 3-hydroxybutanoate fragment
ticarcillin
-
formation of a covalent comple that undergoes non-hydrolytic fragmentation
ZINC02475683
simulation of complex with isoform IprQ. The DELTAG# for the acylation is calculated as 28.26 kcal/mol
ZINC03784242
simulation of complex with isoform IprQ. The DELTAG# for the acylation is calculated as 27.86 kcal/mol
ZINC03788344
simulation of complex with isoform IprQ. The DELTAG# for the acylation is calculated as 13.67 kcal/mol
ZINC03791246
simulation of complex with isoform IprQ. The DELTAG# for the acylation is calculated as 22.88 kcal/mol
Imipenem
irreversible inhibition, imipenem binds only the catalytically active enzyme with apparent submicromolar affinity
Imipenem
investigation of the binding interactions using a two-layered ONIOM model
Imipenem
modeling of the reaction pathways. A 6-membered ring model best describes the inhibition mechanism of acylation. The electrostatic potential and the natural bond orbital analysis show stronger interactions in 6-membered ring transition state mechanism involving water in the active site of the enzyme
meropenem
meropenem binds only the catalytically active enzyme with apparent submicromolar affinity
meropenem
investigation of the binding interactions using a two-layered ONIOM model
meropenem
modeling of the reaction pathways. A 6-membered ring model best describes the inhibition mechanism of acylation. The electrostatic potential and the natural bond orbital analysis show stronger interactions in 6-membered ring transition state mechanism involving water in the active site of the enzyme
(4R,6S)-3-[[(3R,5R)-5-(aminomethyl)oxolan-3-yl]sulfanyl]-6-[(1R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid
compound identified by virtual screening, interacts with residues Arg242 and Gly304
(4R,6S)-3-[[(3R,5R)-5-(aminomethyl)oxolan-3-yl]sulfanyl]-6-[(1R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid
simulation of complex with isoform IprQ. The DELTAG# for the acylation is calculated as 24.29 kcal/mol
2-[(2S,3S)-2-(4-fluorophenyl)-1-(3-methylphenyl)-4-oxoazetidin-3-yl]-5-nitro-1H-isoindole-1,3(2H)-dione
compound identified by virtual screening, interacts with the residue Asn263 and a water molecule which is within the active site
2-[(2S,3S)-2-(4-fluorophenyl)-1-(3-methylphenyl)-4-oxoazetidin-3-yl]-5-nitro-1H-isoindole-1,3(2H)-dione
simulation of complex with isoform IprQ. The DELTAG# for the acylation is calculated as 20.9 kcal/mol
cefotaxime
-
formation of a covalent comple that undergoes non-hydrolytic fragmentation, resulting in an adduct in which the C3' leaving group is lost
cephalothin
-
formation of a covalent comple that undergoes non-hydrolytic fragmentation, resulting in an adduct in which the C3' leaving group is lost
faropenem
-
formation of a covalent comple that undergoes non-hydrolytic fragmentation, yielding LdtB acylated with a 3-hydroxybutanoate fragment
faropenem
high degree of Cys354 modification, fast degradation following the initial acylation event
piperacillin
-
formation of a covalent comple that undergoes non-hydrolytic fragmentation. Cleavage of the C5-C6 bond of the penicillin-derived LdtB acyl-enzyme complex yields LdtB modified by an acetyl group bearing the penicillin C6 side chain
additional information
investigation of the binding interactions carbapenems (biapenem, imipenem, meropenem, and tebipenem) using a two-layered ONIOM model. The carbapenems exhibit reasonable binding interactions towards LdtB. Increasing the number of amino acid residues that form hydrogen bond interactions in the QM layer shows significant impact in binding interaction energy differences and the stabilities of the carbapenems inside the active pocket. The hydrogen bond interactions and charge transfer from the bonding to anti-bonding orbitals between catalytic residues of the enzyme and selected ligands enhances the binding and stability of carbapenem-LdtB complexes
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additional information
computation of the ring strain energy, force constant of amide, acylation transition states and second-order perturbation stabilization energies of 13 basic structural units of beta-lactam derivatives. The calculated DELTAG# values from the acylation reaction of the lactams reveal a faster rate of C-N cleavage in the five-membered ring lactams especially in the 1-2 derivatives. Electronic factors may play more of a role on reactivity of the beta-lactam ring, than ring strain
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additional information
the intact catalytic site is required for the acylation by beta-lactams
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additional information
virtual screening of compounds from the ZINC database against Ldt5 investigated with AutoDock Vina and Schrödinger Maestro software programs. A set of compounds from four antibiotic classes with less than 30 kcal/mol molecular mechanics/generalized born surface area binding free energies is characterized
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additional information
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virtual screening of compounds from the ZINC database against Ldt5 investigated with AutoDock Vina and Schrödinger Maestro software programs. A set of compounds from four antibiotic classes with less than 30 kcal/mol molecular mechanics/generalized born surface area binding free energies is characterized
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
0.65
truncated protein, residues A55-A408, pH 7.5, temperature not specified in the publication
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
?
x * 40900, SDS-PAGE, truncated protein residues A55-A408, including His-tag
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
hanging drop vapor diffusion method, using 0.1 M HEPES (pH 7.5), 1 M succinic acid, and 1% (w/v) PEG MME 2000
5,5'-dithio-bis-2-nitrobenzoic acid derivative, to 1.55 A resolution. The compound binds to the catalytic cysteine residue, Cys354, in both chains of the asymmetric unit
apo and faropenem-acylated forms of Ldt3 at 1.3 and 1.8 A resolution, respectively. The structures revealed a fold and catalytic diad similar to those of other Ldt enzymes. Docking of beta-lactam antibiotics at the active site suggests interaction with conserved amino acids. Faropenem may be degraded after Cys246 acylation, and possibly only a beta-hydroxybutanoate or an acetyl group covalently attached to the enzyme remains
crystal structures of L,D-transpeptidase 2 complexed with biapenem or tebipenem. Biapenem and tebipenem bind to the outer cavity, covalently inactivate the enzyme, and subsequently degrade via an S-conjugate elimination mechanism
structure of L,D-transpeptidase 2 in the apo form and in complex with meropenem and imipenem. The periplasmic region of L,D-transpeptidase 2 folds into three domains. The catalytic residues are situated in the C-terminal domain. The acylation reaction occurs between carbapenem antibiotics and the catalytic Cys354 forming a covalent complex. This adduct formation mimics the acylation of L,D-transpeptidase 2 with the donor PG-stem. In the crystal structures of the apo and the carbapenem complexes, the N-terminal domain has a muropeptide unit non-covalently bound to it
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
C354A
mutant does not display adduct formation with inhibitors faropenem or 6-aminopenicillanic acid
H336A
no cysteine acylation by inhibitors can be detected in a 0-500-s time frame
H352A
mutation does not show a decrease in the rate of the acylation reaction by inhibitors
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
in presence of beta-lactam antibiotics, a thermal shift ranging from -6.1°C for biapenem to 2.2°C for faropenem is observed
additional information
-
in presence of beta-lactam antibiotics, a thermal shift ranging from -6.1°C for biapenem to 2.2°C for faropenem is observed
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
enzyme is sensitive to SH group oxidation. Addition of DTT to the culture medium and enzyme isolation under reducing conditions increases the specific activity activity of the preparation about 2fold
755739
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
nickel affinity column chromatography and Sepharose 12 column chromatography
Ni-NTA agarose resin column chromatography and Superdex 75 gel filtration
-
Ni-NTA agarose resin column chromatography, MonoQ column chromatography, and Superdex HR10/30 gel filtration
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
the extramembrane portion of the L,D-transpeptidase is expressed in Escherichia coli BL21(DE3) cells
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Dubee, V.; Triboulet, S.; Mainardi, J.L.; Etheve-Quelquejeu, M.; Gutmann, L.; Marie, A.; Dubost, L.; Hugonnet, J.E.; Arthur, M.
Inactivation of Mycobacterium tuberculosis L,D-transpeptidase LdtMt1 by carbapenems and cephalosporins
Antimicrob. Agents Chemother.
56
4189-4195
2012
Mycobacterium tuberculosis, Mycobacterium tuberculosis H37Rv
Gupta, R.; Lavollay, M.; Mainardi, J.L.; Arthur, M.; Bishai, W.R.; Lamichhane, G.
The Mycobacterium tuberculosis protein LdtMt2 is a nonclassical transpeptidase required for virulence and resistance to amoxicillin
Nat. Med.
16
466-469
2010
Mycobacterium tuberculosis, Mycobacterium tuberculosis CDC 1551
Erdemli, S.B.; Gupta, R.; Bishai, W.R.; Lamichhane, G.; Amzel, L.M.; Bianchet, M.A.
Targeting the cell wall of Mycobacterium tuberculosis: structure and mechanism of L,D-transpeptidase 2
Structure
20
2103-2115
2012
Mycobacterium tuberculosis (O53223), Mycobacterium tuberculosis
Gokulan, K.; Khare, S.; Cerniglia, C.E.; Foley, S.L.; Varughese, K.I.
Structure and inhibitor specificity of L,D-transpeptidase (LdtMt2) from Mycobacterium tuberculosis and antibiotic resistance calcium binding promotes dimer formation
AAPS J.
20
44
2018
Mycobacterium tuberculosis (I6Y9J2), Mycobacterium tuberculosis, Mycobacterium tuberculosis H37Rv (I6Y9J2)
Libreros-Zuniga, G.A.; Dos Santos Silva, C.; Salgado Ferreira, R.; Dias, M.V.B.
Structural basis for the interaction and processing of beta-lactam antibiotics by L,D-transpeptidase 3 (LdtMt3) from Mycobacterium tuberculosis
ACS Infect. Dis.
5
260-271
2019
Mycobacterium tuberculosis (O06825), Mycobacterium tuberculosis, Mycobacterium tuberculosis H37Rv (O06825)
Baldin, S.; Shcherbakova, T.; Svedas, V.
Isolation, purification and characterization of L,D-transpeptidase 2 from Mycobacterium tuberculosis
Acta Naturae
11
23-28
2019
Mycobacterium tuberculosis (I6Y9J2)
Lohans, C.; Chan, H.; Malla, T.; Kumar, K.; Kamps, J.; McArdle, D.; van Groesen, E.; de Munnik, M.; Tooke, C.; Spencer, J.; Paton, R.; Brem, J.; Schofield, C.
Non-hydrolytic beta-lactam antibiotic fragmentation by L,D-transpeptidases and serine beta-lactamase cysteine variants
Angew. Chem. Int. Ed. Engl.
58
1990-1994
2019
Mycobacterium tuberculosis
Wang, X.; Gu, X.; Zhang, C.; Zhao, F.; Deng, K.
Structural and biochemical analyses of the LdtMt2-panipenem adduct provide new insights into the effect of the 1-beta-methyl group on carbapenems
Biochem. Biophys. Res. Commun.
523
6-9
2020
Mycobacterium tuberculosis (I6Y9J2), Mycobacterium tuberculosis, Mycobacterium tuberculosis ATCC 25618 (I6Y9J2)
Bianchet, M.A.; Pan, Y.H.; Basta, L.A.B.; Saavedra, H.; Lloyd, E.P.; Kumar, P.; Mattoo, R.; Townsend, C.A.; Lamichhane, G.
Structural insight into the inactivation of Mycobacterium tuberculosis non-classical transpeptidase LdtMt2 by biapenem and tebipenem
BMC Biochem.
18
008
2017
Mycobacterium tuberculosis (I6Y9J2), Mycobacterium tuberculosis H37Rv (I6Y9J2)
Tolufashe, G.; Halder, A.; Ibeji, C.; Lawal, M.; Ntombela, T.; Govender, T.; Maguire, G.; Lamichhane, G.; Kruger, H.; Honarparvar, B.
Inhibition of Mycobacterium tuberculosis L,D-transpeptidase 5 by carbapenems MD and QM/MM mechanistic studies
ChemistrySelect
3
13603-13612
2018
Mycobacterium tuberculosis (A5TZL1)
-
Ibeji, C.U.; Lawal, M.M.; Tolufashe, G.F.; Govender, T.; Naicker, T.; Maguire, G.E.M.; Lamichhane, G.; Kruger, H.G.; Honarparvar, B.
The driving force for the acylation of beta-lactam antibiotics by L,D-transpeptidase 2 quantum mechanics/molecular mechanics (QM/MM) study
Chemphyschem
20
1126-1134
2019
Mycobacterium tuberculosis (I6Y9J2), Mycobacterium tuberculosis ATCC 25618 (I6Y9J2)
Steiner, E.M.; Schneider, G.; Schnell, R.
Binding and processing of beta-lactam antibiotics by the transpeptidase LdtMt2 from Mycobacterium tuberculosis
FEBS J.
284
725-741
2017
Mycobacterium tuberculosis (I6Y9J2), Mycobacterium tuberculosis H37Rv (I6Y9J2)
Ntombela, T.; Fakhar, Z.; Ibeji, C.U.; Govender, T.; Maguire, G.E.M.; Lamichhane, G.; Kruger, H.G.; Honarparvar, B.
Molecular insight on the non-covalent interactions between carbapenems and L,D-transpeptidase 2 from Mycobacterium tuberculosis ONIOM study
J. Comput. Aided Mol. Des.
32
687-701
2018
Mycobacterium tuberculosis (O53223), Mycobacterium tuberculosis CDC 1551 (O53223)
Sabe, V.T.; Tolufashe, G.F.; Ibeji, C.U.; Maseko, S.B.; Govender, T.; Maguire, G.E.M.; Lamichhane, G.; Honarparvar, B.; Kruger, H.G.
Identification of potent L,D-transpeptidase 5 inhibitors for Mycobacterium tuberculosis as potential anti-TB leads virtual screening and molecular dynamics simulations
J. Mol. Model.
25
328
2019
Mycobacterium tuberculosis (P9WKV2), Mycobacterium tuberculosis, Mycobacterium tuberculosis CDC 1551 (P9WKV2)
Ibeji, C.U.; Tolufashe, G.F.; Ntombela, T.; Govender, T.; Maguire, G.E.M.; Lamichhane, G.; Kruger, H.G.; Honarparvar, B.
The catalytic role of water in the binding site of l,d-transpeptidase 2 within acylation mechanism A QM/MM (ONIOM) modelling
Tuberculosis
113
222-230
2018
Mycobacterium tuberculosis (O53223), Mycobacterium tuberculosis, Mycobacterium tuberculosis CDC1551 (O53223)
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