Information on EC 2.3.1.122 - trehalose O-mycolyltransferase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.3.1.122
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RECOMMENDED NAME
GeneOntology No.
trehalose O-mycolyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
2 alpha,alpha-trehalose 6-mycolate = alpha,alpha-trehalose + alpha,alpha-trehalose 6,6'-bismycolate
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acyl group transfer
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
mycolate biosynthesis
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SYSTEMATIC NAME
IUBMB Comments
alpha,alpha-trehalose-6-mycolate:alpha,alpha-trehalose-6-mycolate 6'-mycolyltransferase
Catalyses the exchange of mycolic acid between trehalose, trehalose mycolate and trehalose bismycolate. Trehalose 6-palmitate can also act as donor.
CAS REGISTRY NUMBER
COMMENTARY hide
111694-11-2
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain BCG
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
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members of the Ag85 family, Ag85A, Ag85B, and Ag85C, share high sequence and structural homology characterized by an alpha,beta-hydrolase fold and a hydrophobic fibronectin-binding domain. Their active sites are highly conserved, featuring a histidine, aspartic acid or glutamic acid and serine catalytic triad, a hydrophobic tunnel for the lipids and two trehalose binding sites
malfunction
metabolism
glucose causes Mycobacterium avium to down-regulate trehalose dimycolate expression while up-regulating glucose monomycolate. In vitro, the mechanism of reciprocal regulation of trehalose dimycolate and glucose monomycolate involves competitive substrate selection by antigen 85A. The switch from trehalose dimycolate to glucose monomycolate biosynthesis occurs near the physiological concentration of glucose present in mammalian hosts. Furthermore, it is demonstrated that glucose monomycolate is produced in vivo by mcobacteria in mouse lung
physiological function
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
2 alpha,alpha-trehalose 6-monomycolate
alpha,alpha-trehalose 6,6'-dimycolate + alpha,alpha-trehalose
show the reaction diagram
2 alpha,alpha-trehalose 6-mycolate
alpha,alpha-trehalose + alpha,alpha-trehalose 6,6'-bismycolate
show the reaction diagram
alpha,alpha'-trehalose 6-monomycolate + alpha,alpha'-trehalose 6-monomycolate
alpha,alpha'-trehalose 6,6'-dimycolate + alpha,alpha'-trehalose
show the reaction diagram
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-
?
alpha,alpha-trehalose 6,6'-dimycolate + PorA
alpha,alpha-trehalose 6-monomycolate + mycoloyl-PorA
show the reaction diagram
alpha,alpha-trehalose 6,6'-dimycolate + PorH
alpha,alpha-trehalose 6-monomycolate + mycoloyl-PorH
show the reaction diagram
alpha,alpha-trehalose 6-mycolate
alpha,alpha-trehalose + alpha,alpha-trehalose 6,6'-bismycolate
show the reaction diagram
alpha,alpha-trehalose 6-mycolate + alpha,alpha-trehalose
alpha,alpha-trehalose + alpha,alpha-trehalose 6-mycolate
show the reaction diagram
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r
alpha,alpha-trehalose 6-palmitate + trehalose
alpha,alpha-trehalose + alpha,alpha-trehalose 6-palmitate
show the reaction diagram
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r
alpha,alpha-trehalose monomycolate + D-glucose
D-glucose monomycolate + alpha,alpha-trehalose
show the reaction diagram
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
2 alpha,alpha-trehalose 6-mycolate
alpha,alpha-trehalose + alpha,alpha-trehalose 6,6'-bismycolate
show the reaction diagram
alpha,alpha-trehalose 6-mycolate
alpha,alpha-trehalose + alpha,alpha-trehalose 6,6'-bismycolate
show the reaction diagram
alpha,alpha-trehalose 6-mycolate + alpha,alpha-trehalose
alpha,alpha-trehalose + alpha,alpha-trehalose 6-mycolate
show the reaction diagram
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r
additional information
?
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the enzyme, in form of DNA and protein or peptide fragments, is immunogenic in C57BL/6 mice and effective as tuberculosis vaccine against Mycobacterium tuberculosis strain H37Rv, alone and especially in combination with PstS3 DNA and/or protein, detailed overview
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
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no requirement of cofactors such as ATP or CoA
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INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)-methyl ethyl butylphosphonate
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IC50: 0.010 mM
(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)-methyl ethyl tetradecylphosphonate
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IC50: 0.0410 mM
(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)methyl ethyl heptylphosphonate
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IC50: 0.0013 mM
(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)methyl ethyl nonylphosphonate
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IC50: 0.0871 mM
1-(3-phenoxy)benzyl 1-octanesulfonate
IC50: 0.0043 mM
2-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)-ethyl ethyl butylphosphonate
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IC50: 0.0507 mM
2-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)-ethyl ethyl nonylphosphonate
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IC50: 0.0257 mM
2-amino-6-propyl-4,5,6,7-tetrahydro-1-benzothiophene-3-carbonitrile
i.e. I3-AG85, enzyme inhibition leads to accumulation of trehalose monomycolate and disruption of the bacterial envelope, I3-AG85 also inhibits Mycobacterium tuberculosis survival in infected primary macrophages. Binding of I3-AG85 to Ag85C is similar to its binding to the artificial substrate octylthioglucoside, overview
2-phenylethanol
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3,4,5-trimethoxybenzylphosphonate
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IC50: 0.102 mM
3-Phenoxybenzyl alcohol
trehalose mimetic inhibitor
4-(phthalimido)butanol
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6-azido-6-deoxytrehalose
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inhibits all three members of Ag85 complex in vitro
ACP
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0.00056 mg/0.6 ml, 68% reduction of activity
ethyl 3-phenoxybenzyl butylphosphonate
ethyl 3-phenoxybenzyl heptylphosphonate
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IC50: 0.021 mM
ethyl 3-phenoxybenzyl hexylphosphonate
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IC50: 0.0426 mM
ethyl 3-phenoxybenzyl nonylphosphonate
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IC50: 0.0148 mM
ethyl 3-phenoxybenzyl tetradecylphosphonate
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IC50: 0.862 mM
ethyl hydrogen 2-phenylethyl phosphonate
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IC50: 0.225 mM
ethyl hydrogen 4-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)butylphosphonate
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IC50: 0.110 mM
ethyl hydrogen butylphosphonate
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IC50: 0.431 mM
ethyl hydrogen heptylphosphonate
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IC50: 0.0011 mM
ethyl hydrogen hexylphosphonate
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IC50: 0.0036 mM
ethyl hydrogen nonylphosphonate
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IC50: 0.471 mM
iodoacetamide
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16.7 mM, 53% decrease of activity
MgCl2
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16 mM, 91% inhibition
N-(hydroxyethyl)phthalimide
trehalose mimetic inhibitor
N-(omega-hydroxyalky)phthalimide
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Tween 80
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strong, might disrupt necessary structure of the TM vesicle/micelle
additional information
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.043 - 0.459
alpha,alpha-trehalose 6-monomycolate
0.0083
trehalose
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.09
alpha,alpha-trehalose 6-monomycolate
Mycobacterium smegmatis
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pH 5.7, 37°C
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.01
(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)-methyl ethyl butylphosphonate
Mycobacterium avium
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IC50: 0.010 mM
0.041
(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)-methyl ethyl tetradecylphosphonate
Mycobacterium avium
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IC50: 0.0410 mM
0.0013
(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)methyl ethyl heptylphosphonate
Mycobacterium avium
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IC50: 0.0013 mM
0.0871
(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)methyl ethyl nonylphosphonate
Mycobacterium avium
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IC50: 0.0871 mM
0.0043
1-(3-phenoxy)benzyl 1-octanesulfonate
Mycobacterium tuberculosis
P9WQN9
IC50: 0.0043 mM
0.0507
2-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)-ethyl ethyl butylphosphonate
Mycobacterium avium
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IC50: 0.0507 mM
0.0257
2-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)-ethyl ethyl nonylphosphonate
Mycobacterium avium
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IC50: 0.0257 mM
0.102
3,4,5-trimethoxybenzylphosphonate
Mycobacterium avium
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IC50: 0.102 mM
0.002
ethyl 3-phenoxybenzyl butylphosphonate
0.021
ethyl 3-phenoxybenzyl heptylphosphonate
Mycobacterium avium
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IC50: 0.021 mM
0.0426
ethyl 3-phenoxybenzyl hexylphosphonate
Mycobacterium avium
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IC50: 0.0426 mM
0.0148
ethyl 3-phenoxybenzyl nonylphosphonate
Mycobacterium avium
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IC50: 0.0148 mM
0.862
ethyl 3-phenoxybenzyl tetradecylphosphonate
Mycobacterium avium
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IC50: 0.862 mM
0.225
ethyl hydrogen 2-phenylethyl phosphonate
Mycobacterium avium
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IC50: 0.225 mM
0.11
ethyl hydrogen 4-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)butylphosphonate
Mycobacterium avium
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IC50: 0.110 mM
0.431
ethyl hydrogen butylphosphonate
Mycobacterium avium
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IC50: 0.431 mM
0.0011
ethyl hydrogen heptylphosphonate
Mycobacterium avium
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IC50: 0.0011 mM
0.0036
ethyl hydrogen hexylphosphonate
Mycobacterium avium
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IC50: 0.0036 mM
0.471
ethyl hydrogen nonylphosphonate
Mycobacterium avium
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IC50: 0.471 mM
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 9
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45% of maximal activity at pH 5.0, 30% of maximal activity at pH 9.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25 - 55
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75% of maximal activity at 25°C, 12% at 55°C
PDB
SCOP
CATH
ORGANISM
UNIPROT
Corynebacterium glutamicum (strain ATCC 13032 / DSM 20300 / JCM 1318 / LMG 3730 / NCIMB 10025)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25000
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SDS-PAGE, gel filtration
30000
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FbpA: SDS-PAGE, Western Blot, aminoacid sequence. FbpB: aminoacid sequence. FbpC2: SDS-PAGE, Western Blot, aminoacid sequence
32000
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SDS-PAGE
33000
SDS-PAGE, detection of a fusion protein consiting of a His-tag and the mature Ag85A protein
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
three-dimensional model based on crystal structure of related enzymes, comparison of enzymes
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native protein, to 1.75 A resolution. Strucutre shows the presence of a unique loop that is absent from enzymes that transfer mycoloyl residues onto both trehalose and the cell wall arabinogalactan
three-dimensional model based on crystal structure of related enzymes, comparison of enzymes
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GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
buffer of ionic strength of 0.1 M and above stabilizes
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dithiothreitol, 1 mM, stabilizes
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purification has to be carried out at 4°C
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate fractionation, gel filtration and Sephadex column chromatography
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native enzyme from strain BCG, recombinant enzyme from Escherichia coli
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recombinant proteins purified by affinity chromatography on Ni2+ charged His-trap columns
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using Ni-NTA chromatography
using Ni-NTA chromatography and by applying a linear imidazole gradient from 10 to 500 mM. The final purification step contains gel filtration on a Superdex 200 column
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed as a His-tagged fusion protein
expressed in Escherichia coli as a His-tagged fusion protein
expression of Ag85A in baby hamster kidney cells and as His-tagged enzyme in Escherichia coli
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expression of FbpA, FbpB and FbpC2 in Escherichia coli, FbpB poorly expressed due to differential efficiency of mRNA translation
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recombinant expression as C-terminally His-tagged enzyme
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
S130L
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mutant S130L of Mycobacterium avium exhibits a reduced mycosyltransferase activity compared to wild-type
L130S
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leucine at position 130 of Mycobacterium leprae FbpA limits access of mycolate-containing glycolipid substrates to their binding site. Mutant L130S exhibits an enhanced mycosyltransferase activity compared to wild-type while the ability to transfer the unbranched acyl chain is unchanged. Km (substrate: short-chain trehalose monomycolate): 0.043 mM, sharply decreased compared to wild-type
additional information
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deletion mutant of enzyme genes cmytA and cmytB. Double mutant fails to grow at 34°C. It exhibits a growth defect and forms segmentation-defective cells at 30°C. Double mutant elaborates 60% less cell-wall-bound corynomycolates and produces less crystalline surface layer proteins than wild-type
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
biotechnology
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an enzyme assay using the natural substrate trehalose dimycolate is developed. The colorimetric assay is based on the quantification of glucose from the degradation of trehalose, which is the product from catalytic activity of antigen 85A. This assay is suitable for robust high-throughput screening (HTS) for compound library screening against mycolyltransferase. The assay has a very low coefficient of variance (0.04) in 96-well plates and shows a Z' factor of 0.67-0.73, indicating the robustness of the assay
drug development
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antigen 85 proteins are potential drug targets
medicine
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the enzyme is immunogenic in C57BL/6 mice and effective as tuberculosis vaccine, usage of vaccines combining mycolyl-transferase Ag85A and phosphate transport receptor PstS-3, overview
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