Information on EC 2.7.1.136 - macrolide 2'-kinase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.7.1.136
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RECOMMENDED NAME
GeneOntology No.
macrolide 2'-kinase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + oleandomycin = ADP + oleandomycin 2'-O-phosphate
show the reaction diagram
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phospho group transfer
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-
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SYSTEMATIC NAME
IUBMB Comments
ATP:macrolide 2'-O-phosphotransferase
Erythromycin, spiramycin and some other macrolide antibiotics can also act as acceptors.
CAS REGISTRY NUMBER
COMMENTARY hide
116036-69-2
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain CU1, gene mphB
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Manually annotated by BRENDA team
strain UC 5240
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Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + azithromycin
ADP + azithromycin 2'-O-phosphate
show the reaction diagram
ATP + clarithromycin
?
show the reaction diagram
ATP + clarithromycin
ADP + clarithromycin 2'-O-phosphate
show the reaction diagram
ATP + erythromycin
ADP + anhydroerythromycin-2'-O-phosphate
show the reaction diagram
ATP + erythromycin
ADP + erythromycin 2'-O-phosphate
show the reaction diagram
ATP + josamycin
?
show the reaction diagram
ATP + josamycin
ADP + josamycin 2'-O-phosphate
show the reaction diagram
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15-membered ring macrolide
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-
?
ATP + kitasamycin
?
show the reaction diagram
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52% activity with respect to oleandomycin for strain CU1, 61% activity with respect to oleandomycin for strain BM 2506
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-
?
ATP + kitasamycin
ADP + kitasamycin 2'-O-phosphate
show the reaction diagram
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15-membered ring macrolide
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-
?
ATP + midecamycin
?
show the reaction diagram
ATP + neomycin
ADP + ?
show the reaction diagram
-
-
-
?
ATP + oleandomycin
ADP + oleandomycin 2'-O-phosphate
show the reaction diagram
ATP + rokitamycin
ADP + rokitamycin 2'-O-phosphate
show the reaction diagram
-
15-membered ring macrolide
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-
?
ATP + roxithromycin
?
show the reaction diagram
ATP + roxithromycin
ADP + roxithromycin 2'-O-phosphate
show the reaction diagram
ATP + spectinomycin
?
show the reaction diagram
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5% activity with respect to oleandomycin
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-
?
ATP + spiramycin
ADP + spiramycin 2'-O-phosphate
show the reaction diagram
ATP + telithromycin
ADP + telithromycin 2'-O-phosphate
show the reaction diagram
ATP + triacetyloleandomycin
?
show the reaction diagram
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100% activity with respect to oleandomycin
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-
?
ATP + triacetyloleandomycin
ADP + triacetyloleandomycin 2'-O-phosphate
show the reaction diagram
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as active as oleandomycin
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?
ATP + troleandomycin
?
show the reaction diagram
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82% activity with respect to oleandomycin for strain CU1, 105% activity with respect to oleandomycin for strain BM 2506
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-
?
ATP + troleandomycin
ADP + troleandomycin 2'-O-phosphate
show the reaction diagram
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14-membered ring macrolide
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-
?
ATP + tylosin
ADP + tylosin 2'-O-phosphate
show the reaction diagram
ATP + tylosin
ADP + tylosin 2'-phosphate
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
ATP + azithromycin
ADP + azithromycin 2'-O-phosphate
show the reaction diagram
Q5W3B4
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-
-
?
ATP + clarithromycin
ADP + clarithromycin 2'-O-phosphate
show the reaction diagram
Q5W3B4
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-
-
?
ATP + erythromycin
ADP + anhydroerythromycin-2'-O-phosphate
show the reaction diagram
Q5W3B4
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-
-
?
ATP + roxithromycin
ADP + roxithromycin 2'-O-phosphate
show the reaction diagram
Q5W3B4
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-
-
?
ATP + tylosin
ADP + tylosin 2'-O-phosphate
show the reaction diagram
Q5W3B4
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-
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?
additional information
?
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Q47396
under physiological conditions, enzyme exclusively uses ATP, no substrate: GTP
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
potassium chloride enhances reactivity, it is added in the reaction mixture at 0.06 M
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Adenylyl imidodiphosphate
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CoSO4
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1 mM, 51% inhibition
CuSO4
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1 mM, 52% inhibition
EDTA
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10 mM, 100% inhibition
FeSO4
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1 mM, 66% inhibition
Iodine
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1 mM, 100% inhibition
oleandomycin
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ZnSO4
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1 mM, 93% inhibition
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
erythromycin
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crude cell extracts from cells treated with erythromycin show increased activity
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0106
ATP
pH 7.5, 37C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.59
ATP
Escherichia coli
Q47396
pH 7.5, 37C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
150
ATP
Escherichia coli
Q47396
pH 7.5, 37C
4
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0142
Adenylyl imidodiphosphate
pH 7.5, 37C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0007
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purified recombinant mutant H205A, substrate oleandomycin
0.04
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purified recombinant wild-type enzyme, substrate roxithromycin or rokitamycin
0.05
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purified recombinant wild-type enzyme, substrate josamycin or tylosin
0.052
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purified recombinant mutant H198A, substrate oleandomycin
0.056
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purified recombinant mutant H205N, substrate oleandomycin
0.06
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purified recombinant wild-type enzyme, substrate troleandomycin, clarithromycin, azithromycin, or erythromycin
0.1
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purified recombinant wild-type enzyme, substrate oleandomycin
0.16
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purified recombinant wild-type enzyme, substrate kitasamycin
0.21
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purified recombinant wild-type enzyme, substrate spiramycin
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.8
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assay at
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
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assay at
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
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mph gene is located on nontransmissible plasmid DNA for Escherichia coli strain CU1
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Manually annotated by BRENDA team
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 34000, SDS-PAGE
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, Tris-HCl buffer pH 7, 0.06 M KCl, 0.01 M MgCl2, 0.006 M 2-mercaptoethanol, stable
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant wild-type and mutant enzymes from strain TG1
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli DH5alpha, complete pRSB105 macrolide resistance gene region cloned into the vector pBCSK, recombinant plasmid pNK-mph carries the complete mph gene in pBluescript II KS, transcription from the constructed plasmids analyzed by RT-PCR, constructs tested for conferring erythromycin and azithromycin resistance
expressed in Escherichia coli, strain DH5alpha mcr containing the pBCSK vector carrying a chloramphenicol resistance gene and the pRSB111 erythromycin resistance-conferring region, recombinant expression in Pseudomonas sp. strain B13 GFP1
expression of wild-type and mutant enzymes in strains TG1 and N43
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mph gene for Escherichia coli strain CU1 is identical to that previously reported for mphB from Escherichia coli strain BM 2506
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mph genes cloned into Escherichia coli AG100A
mph genes cloned into Escherichia coli AG100A; mph genes cloned into Escherichia coli AG100A
mphA gene is arranged in close proximity to mrx and MphR(A) which are proteins probably involved in mph regulation, they all form a gene cluster
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
H198A
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site-directed mutagenesis, 50% reduction of the minimal inhibitory concentration MIC level of oleandomycin and 50% reduced activity compared to the wild-type enzyme
H205A
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site-directed mutagenesis, 87.5% reduction of the minimal inhibitory concentration MIC level of oleandomycin and 50% reduced activity compared to the wild-type enzyme
H205N
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site-directed mutagenesis, unaffected minimal inhibitory concentration MIC level of oleandomycin, but more than 99% reduced activity compared to the wild-type enzyme
H198A
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site-directed mutagenesis, 50% reduction of the minimal inhibitory concentration MIC level of oleandomycin and 50% reduced activity compared to the wild-type enzyme
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H205A
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site-directed mutagenesis, 87.5% reduction of the minimal inhibitory concentration MIC level of oleandomycin and 50% reduced activity compared to the wild-type enzyme
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H205N
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site-directed mutagenesis, unaffected minimal inhibitory concentration MIC level of oleandomycin, but more than 99% reduced activity compared to the wild-type enzyme
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
competitive assay that mimics in vivo nucleotide triphosphate concentrations and usage by the enzyme. Downstream analysis of reaction products by high-performance liquid chromatography enables the determination of partitioning of phosphate flux from nucleotide triphosphate donors to antibiotics