Information on EC 2.7.1.197 - protein-Npi-phosphohistidine-D-mannitol phosphotransferase

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The expected taxonomic range for this enzyme is: Escherichia coli

EC NUMBER
COMMENTARY hide
2.7.1.197
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RECOMMENDED NAME
GeneOntology No.
protein-Npi-phosphohistidine-D-mannitol phosphotransferase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
[protein]-Npi-phospho-L-histidine + D-mannitol[side 1] = [protein]-L-histidine + D-mannitol 1-phosphate[side 2]
show the reaction diagram
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Fructose and mannose metabolism
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SYSTEMATIC NAME
IUBMB Comments
protein-Npi-phospho-L-histidine:D-mannitol Npi-phosphotransferase
This enzyme is a component (known as enzyme II) of a phosphoenolpyruvate (PEP)-dependent, sugar transporting phosphotransferase system (PTS). The system, which is found only in prokaryotes, simultaneously transports its substrate from the periplasm or extracellular space into the cytoplasm and phosphorylates it. The phosphate donor, which is shared among the different systems, is a phospho-carrier protein of low molecular mass that has been phosphorylated by EC 2.7.3.9 (phosphoenolpyruvate---protein phosphotransferase). Enzyme II, on the other hand, is specific for a particular substrate, although in some cases alternative substrates can be transported with lower efficiency. The reaction involves a successive transfer of the phosphate group to several amino acids within the enzyme before the final transfer to the substrate.
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
[heat stable phosphocarrier protein HPr]-Npi-phospho-L-histidine + D-mannitol[side 1]
[heat stable phosphocarrier protein HPr]-L-histidine + D-mannitol 1-phosphate[side 2]
show the reaction diagram
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?
[heat-stable phosphocarrier protein]-Npi-phospho-L-histidine + D-mannitol[side 1]
[heat-stable phosphocarrier protein]-L-histidine + D-mannitol 1-phosphate[side 2]
show the reaction diagram
[protein HPr]-Npi-phospho-L-histidine + D-mannitol[side 1]
[protein HPr]-L-histidine + D-mannitol 1-phosphate[side 2]
show the reaction diagram
[protein]-L-histidine + D-mannitol 1-phosphate[side 2]
[protein]-Npi-phospho-L-histidine + D-mannitol[side 1]
show the reaction diagram
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r
[protein]-Npi-phospho-L-histidine + 2-amino-2-deoxy-D-mannitol[side 1]
[protein]-L-histidine + 2-amino-2-deoxy-D-mannitol 1-phosphate[side 2]
show the reaction diagram
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50% activity compared to D-mannitol
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-
?
[protein]-Npi-phospho-L-histidine + 2-deoxy-D-mannitol[side 1]
[protein]-L-histidine + 2-deoxy-D-mannitol 1-phosphate[side 2]
show the reaction diagram
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32% activity compared to D-mannitol
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-
?
[protein]-Npi-phospho-L-histidine + D-arabitol[side 1]
[protein]-L-histidine + D-arabitol 1-phosphate[side 2]
show the reaction diagram
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11% activity compared to D-mannitol
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-
?
[protein]-Npi-phospho-L-histidine + D-glucitol[side 1]
[protein]-L-histidine + D-glucitol 1-phosphate[side 2]
show the reaction diagram
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37% activity compared to D-mannitol
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-
?
[protein]-Npi-phospho-L-histidine + D-mannitol[side 1]
[protein]-L-histidine + D-mannitol 1-phosphate[side 2]
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
[heat stable phosphocarrier protein HPr]-Npi-phospho-L-histidine + D-mannitol[side 1]
[heat stable phosphocarrier protein HPr]-L-histidine + D-mannitol 1-phosphate[side 2]
show the reaction diagram
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-
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?
[heat-stable phosphocarrier protein]-Npi-phospho-L-histidine + D-mannitol[side 1]
[heat-stable phosphocarrier protein]-L-histidine + D-mannitol 1-phosphate[side 2]
show the reaction diagram
[protein HPr]-Npi-phospho-L-histidine + D-mannitol[side 1]
[protein HPr]-L-histidine + D-mannitol 1-phosphate[side 2]
show the reaction diagram
[protein]-L-histidine + D-mannitol 1-phosphate[side 2]
[protein]-Npi-phospho-L-histidine + D-mannitol[side 1]
show the reaction diagram
P00550
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r
[protein]-Npi-phospho-L-histidine + D-mannitol[side 1]
[protein]-L-histidine + D-mannitol 1-phosphate[side 2]
show the reaction diagram
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Mg2+
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Mg2+ ions bind specifically to enzyme, inducing dimerization
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2-amino-2-deoxy-D-mannitol
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63% residual activity at 2 mM
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2-deoxy-D-mannitol
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73% residual activity at 2 mM; 74% residual activity at 2 mM
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D-arabitol
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D-fructose 1,6-bisphosphate
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34% inhibition at 20 mM
D-glucitol
D-glucose 1-phosphate
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D-glucose 6-phosphate
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D-mannitol
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strong substrate inhibition at pH 7.0
D-mannitol 1-phosphate
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strong substrate inhibition at pH 7.0
D-mannoheptitol
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9% residual activity at 2 mM
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D-mannonic acid
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diethyldicarbonate
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complete inactivation at 1 mM
Mg2+
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Mg2+ ions at sufficiently high concentrations (5 mM) inhibit enzyme
N-ethylmaleimide
phosphate
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a 5fold dilution of a mixture of 10 mM phosphate, 5 mM MgSO4, and 50 mM NaF into phosphate buffers, pH 6.0-6.3 just before the zero time point leads to a rapid inhibition of the enzyme activity
Trypsin
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additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
cardiolipin
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phosphate
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inorganic phosphate increases the Vmax, of the transphosphorylation reaction 4.5-fold and increases Km for both substrates approximately 2fold
phosphatidylethanolamine
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phosphatidylglycerol
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additional information
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the purified enzyme is dependent on lubrol PX and phospholipid for maximal activity
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.2
2-amino-2-deoxy-D-mannitol[side 1]
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at pH 8.0 and 37C
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0.5
2-deoxy-D-mannitol[side 1]
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0.066 - 0.29
D-mannitol 1-phosphate[side 2]
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0.01 - 0.46
D-mannitol[side 1]
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.023
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unpurified enzyme from membrane, at pH 9.1 and 37C
26.67
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after 235fold purification, at pH 9.1 and 37C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60000 - 65000
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gel filtration
60000
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gel filtration
65000
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gel filtration
130000
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gel filtration
315000
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globular protein solubilized in decylPEG detergent, gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
heterodimer
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1 * 34000 + 1 * 29000, SDS-PAGE
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
phosphoprotein
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
mannitol-specific IIA domain in complex with HPr protein, vapor diffusion method, using 36-38% (w/v) PEG4000, 0.1 mM N-Tris(hydroxymethyl)methyl-3-aminopropane sulfonic acid/NaOH, pH 8.7, 5 mM Na2EDTA and 1 mM NaN3
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
DEAE-cellulose column chromatography
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DEAE-cellulose column chromatography and hexyl agarose column chromatography
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hexyl agarose column chromatography, omega-aminohexyl agarose column chromatography, and DEAE cellulose column chromatography
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hexylagarose column chromatography and TSK-250 gel filtration
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Q-Sepharose and SP-Sepharose column chromatography
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Q-Sepharose column chromatography and Sephadex G75 gel filtration
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Sephacryl S-200 gel filtration
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli LGS322 cells
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expressed in Escherichia coli strain LGS322
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selenomethionine-labelled enzyme is expressed in Escherichia coli LE392 cells
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C384D
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inactive
C384H
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inactive
E218A
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the mutant has a high Km for mannitol but can carry out facilitated diffusion of mannitol at a rate higher than the wild type protein in its unphosphorylated form
E257A
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the mutant does not detectably bind mannitol up to a concentration of at least 1 mM
G253E
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the mutant cannot transport but still can phosphorylate mannitol
H156A
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the mutant shows wild type activity
H195N
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the mutant shows 30% of wild type activity
H554A
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inactive
H554D
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inactive