Information on EC 2.7.1.190 - aminoglycoside 2''-phosphotransferase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.7.1.190
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RECOMMENDED NAME
GeneOntology No.
aminoglycoside 2''-phosphotransferase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
GTP + gentamicin = GDP + gentamicin 2''-phosphate
show the reaction diagram
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-
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SYSTEMATIC NAME
IUBMB Comments
GTP:gentamicin 2''-O-phosphotransferase
Requires Mg2+. This bacterial enzyme phosphorylates many 4,6-disubstituted aminoglycoside antibiotics that have a hydroxyl group at position 2'', including kanamycin A, kanamycin B, tobramycin, dibekacin, arbekacin, amikacin, gentamicin C, sisomicin and netilmicin. In most, but not all, cases the phosphorylation confers resistance against the antibiotic. Some forms of the enzyme use ATP as a phosphate donor in appreciable amount. The enzyme is often found as a bifunctional enzyme that also catalyses 6'-aminoglycoside N-acetyltransferase activity. The bifunctional enzyme is the most clinically important aminoglycoside-modifying enzyme in Gram-positive bacteria, responsible for high-level resistance in both Enterococci and Staphylococci.
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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
ATP + gentamicin
ADP + gentamicin 2''-phosphate
show the reaction diagram
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GTP is preferred over ATP
-
?
ATP + kanamycin
ADP + kanamycin 2''-phosphate
show the reaction diagram
-
-
-
?
ATP + kanamycin A
ADP + kanamycin A 2''-phosphate
show the reaction diagram
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-
-
-
?
ATP + tobramycin
ADP + tobramycin 2''-phosphate
show the reaction diagram
-
-
-
?
GTP + amikacin
GDP + amikacin 2''-phosphate
show the reaction diagram
GTP + arbekacin
GDP + arbekacin 2''-phosphate
show the reaction diagram
GTP + dibekacin
GDP + dibekacin 2''-phosphate
show the reaction diagram
GTP + gentamicin C
GDP + gentamicin C 2''-phosphate
show the reaction diagram
GTP + isepamicin
GDP + isepamicin 2''-phosphate
show the reaction diagram
-
-
-
?
GTP + kanamycin
GDP + kanamycin 2''-phosphate
show the reaction diagram
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-
-
?
GTP + kanamycin A
GDP + kanamycin A 2''-phosphate
show the reaction diagram
GTP + kanamycin B
GDP + kanamycin B 2''-phosphate
show the reaction diagram
GTP + netilmicin
GDP + netilmicin 2''-phosphate
show the reaction diagram
GTP + sisomicin
GDP + sisomicin 2''-phosphate
show the reaction diagram
GTP + tobramycin
GDP + tobramycin 2''-phosphate
show the reaction diagram
ITP + dibekacin
IDP + dibekacin 2''-phosphate
show the reaction diagram
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-
-
-
?
UTP + tobramycin
UDP + tobramycin 2''-phosphate
show the reaction diagram
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-
-
-
?
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
additional information
?
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P0A0C1
GTP is the exclusive phosphate donor at intracellular nucleotide levels
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INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
apramycin
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-
butirosin
-
-
butirosin A/B
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lividomycin A
neamine
neomycin
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neomycin B
paromomycin
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.033 - 0.26
amikacin
0.0008 - 0.0062
arbekacin
-
0.055 - 2
ATP
0.0003 - 0.0012
dibekacin
0.0007 - 0.006
gentamicin C
0.0008 - 0.168
GTP
0.56
isepamicin
pH 7.0, 22°C
0.039
ITP
-
pH 7.5, 25°C, cosubstrate kanamycin A
0.0011 - 0.0045
kanamycin A
0.00055
kanamycin B
pH 7.0, 22°C
0.0041 - 0.01
netilmicin
0.0008 - 0.0016
sisomicin
0.001
tobramycin
above 0.001, pH 7.0, 22°C
0.1
UTP
-
pH 7.5, 25°C, cosubstrate kanamycin A
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3.7 - 6.5
amikacin
2.3 - 4.3
arbekacin
-
0.09 - 4.37
ATP
0.15 - 0.79
dibekacin
0.5 - 8.3
gentamicin C
0.4 - 5.03
GTP
0.25
isepamicin
Campylobacter jejuni
Q4VR94
pH 7.0, 22°C
2.1
ITP
Enterococcus sp.
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pH 7.5, 25°C, cosubstrate kanamycin A
0.57 - 1.7
kanamycin A
0.12 - 0.38
kanamycin B
0.7 - 18
netilmicin
8.1 - 17
sisomicin
0.11 - 0.7
tobramycin
0.14
UTP
Enterococcus sp.
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pH 7.5, 25°C, cosubstrate kanamycin A
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
25 - 110
amikacin
1394
690 - 2900
arbekacin
206872
0.19 - 63
ATP
4
500 - 2300
dibekacin
6512
1800 - 12000
gentamicin C
15768
23 - 510
GTP
37
0.45
isepamicin
Campylobacter jejuni
Q4VR94
pH 7.0, 22°C
36503
56
ITP
Enterococcus sp.
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pH 7.5, 25°C, cosubstrate kanamycin A
229
180 - 1500
kanamycin A
789
690
kanamycin B
Campylobacter jejuni
Q4VR94
pH 7.0, 22°C
3078
670 - 16000
netilmicin
2604
10000 - 11000
sisomicin
3365
480 - 2200
tobramycin
939
1.4
UTP
Enterococcus sp.
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pH 7.5, 25°C, cosubstrate kanamycin A
65
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.14
butirosin A/B
pH 7.0, 22°C
-
0.011 - 0.012
lividomycin A
0.00016
neamine
pH 7.0, 22°C
0.00026 - 0.00073
neomycin B
0.0053 - 0.0085
paromomycin
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
56000
x * 56000, SDS-PAGE, x * 56850, calculated
56850
x * 56000, SDS-PAGE, x * 56850, calculated
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
apo form and in complex with a bound antibiotic, tobramycin and kanamycin A, to 2.05 A, 1.8 A and 2.15 A resolution, respectively. Substrate binding induces conformational changes, involving rotational shifts of two distinct segments of the enzyme. The helical subdomain is important in substrate specificity
crystal structures of isoform APH(2'')-IVa, wild-type and mutant F95M, in complex with either adenosine or guanosine
to 2.2 A resolution. Access to the ATP-binding template is blocked by a bulky tyrosine residue. Substitution of this tyrosine by a smaller amino acid opens access to the ATP template
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
F95M
no significantly different binding affinities as compared with the wild-type
F95Y
mutation shifts the nucleotide selectivity from a 2.5fold preference for ATP to a 1.5fold preference for GTP
Y92A
residue Y92 blocks access to the ATP-binding template. Mutant shows 8fold decrease in km value for ATP
additional information