Information on EC 1.6.1.1 - NAD(P)+ transhydrogenase (Si-specific)

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The expected taxonomic range for this enzyme is: Bacteria, Eukaryota

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EC NUMBER
COMMENTARY hide
1.6.1.1
-
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RECOMMENDED NAME
GeneOntology No.
NAD(P)+ transhydrogenase (Si-specific)
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
NADPH + NAD+ = NADP+ + NADH
show the reaction diagram
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oxidation
-
-
-
-
redox reaction
-
-
-
-
reduction
-
-
-
-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
NAD/NADH phosphorylation and dephosphorylation
-
-
NAD metabolism
-
-
Nicotinate and nicotinamide metabolism
-
-
Metabolic pathways
-
-
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SYSTEMATIC NAME
IUBMB Comments
NADPH:NAD+ oxidoreductase (Si-specific)
The enzyme from Azotobacter vinelandii is a flavoprotein (FAD). It is Si-specific with respect to both NAD+ and NADP+. Also acts on deamino coenzymes [cf. EC 1.6.1.2 NAD(P)+ transhydrogenase (Re/Si-specific)].
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CAS REGISTRY NUMBER
COMMENTARY hide
9014-18-0
not distinguished from EC 1.6.1.2
9072-60-0
not distinguished from EC 1.6.1.2
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
Azotobacter agilis
-
-
-
Manually annotated by BRENDA team
Chromatium sp.
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
pea
-
-
Manually annotated by BRENDA team
potato tubers
-
-
Manually annotated by BRENDA team
formerly Pseudomonas natriegens
-
-
Manually annotated by BRENDA team
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GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
two pyridine nucleotide transhydrogenases: the energy-independent soluble pyridine nucleotide transhydrogenase (STH or UdhA) (EC 1.6.1.1) and the membrane-bound, energy-dependent pyridine nucleotide transhydrogenase (TH or PntAB) (EC 1.6.1.2). PntAB is widely distributed in the mitochondria and some bacteria, while STH is found only in certain Gammaproteobacteria and gram-positive bacteria
physiological function
the soluble pyridine nucleotide transhydrogenase, STH, is an energy-independent flavoprotein that directly catalyzes hydride transfer between NAD(H) and NADP(H) to maintain homeostasis of these two redox cofactors
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SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
NAD(P)H + 2,6-dichlorophenolindophenol
NAD(P)+ + reduced 2,6-dichlorophenolindophenol
show the reaction diagram
-
-
-
?
NAD(P)H + K4Fe(CN)6
NAD(P)+ + K3Fe(CN)6
show the reaction diagram
-
-
-
?
NADH + 2'-NADP+
NAD+ + 2'-NADPH
show the reaction diagram
-
-
-
?
NADH + 3'-NADP+
NAD+ + 3'-NADPH
show the reaction diagram
-
-
-
?
NADH + NADP+
NADPH + NAD+
show the reaction diagram
-
-
-
?
NADH + thio-NAD+
NAD+ + thio-NADH
show the reaction diagram
-
-
-
?
NADH + thio-NADP+
NAD+ + thio-NADPH
show the reaction diagram
NADP+ + NADH
NADPH + NAD+
show the reaction diagram
NADPH + 3-acetylpyridine-NAD+
NADP+ + 3-acetylpyridine-NADH
show the reaction diagram
NADPH + deamino-NAD+
NADP+ + deamino-NADH
show the reaction diagram
NADPH + NAD+
NADP+ + NADH
show the reaction diagram
NADPH + pyridine aldehyde-NAD+
NADP+ + pyridine aldehyde-NADH
show the reaction diagram
NADPH + thio-NAD+ + H+[side 1]
NADP+ + thio-NADH + H+[side 2]
show the reaction diagram
NADPH + thio-NADP+ + H+[side 1]
NADP+ + thio-NADPH + H+[side 2]
show the reaction diagram
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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
NADH + NADP+
NADPH + NAD+
show the reaction diagram
-
-
-
?
NADPH + NAD+
NADP+ + NADH
show the reaction diagram
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
flavin
flavoprotein
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
EDTA
-
slight activating at low buffer concentrations
K+
-
no full activation compared to Ca2+
Mn2+
-
compared to Ca2+ 10 times higher concetrations are required to obtain the same degree of activation
additional information
not influenced by Na+, Rb+, K+, Li+, and Mg2+
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INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2'-AMP
2-mercaptoethanol
-
5'-AMP
-
5 mM, 92% inhibition of NADP+ reduction, complete inhibition of NAD+ reduction
ADP
-
5 mM, 94% inhibition of NADP+ reduction, complete inhibition of NAD+ reduction
arsenate
-
complete inhibition of activity in either direction
ATP
-
5 mM, 81% inhibition of NADP+ reduction, complete inhibition of NAD+ reduction
beta-mercaptoethanol
72% residual activity at 0.2% (v/v)
Ca2+
91.2% residual activity at 2 mM; slightly inhibitory
CTP
-
5 mM, 86% inhibition of NADP+ reduction
Cu2+
; complete inhibition at 2 mM
deoxycholate
-
-
diphosphate
-
5 mM, 91% inhibition of NADP+ reduction
dithiothreitol
75.3% residual activity at 2 mM
DMSO
slightly inhibitory
EDTA
; 72.6% residual activity at 2 mM
GTP
-
5 mM, 89% inhibition of NADP+ reduction
Mn2+
; 67% residual activity at 2 mM
NAD+
-
competitive to thio-NAD+, uncompetitive with respect to NADPH
NADP+
NADPH
EcSTH activity is strongly inhibited by excess NADPH, but not by thio-NAD+; the enzyme is strongly inhibited by excess NADPH
Ni2+
; 7.4% residual activity at 2 mM
p-Aminophenylarsenoxide
-
0.1 mM, 40-60% inhibition in the absence of either phosphate or magnesium ions, reduction of NAD+ by NADPH in cell-free extracts is rapidly and completely inhibited in the presence of 20 mM phosphate
p-chloromercuribenzoate
-
0.044 mM, 40-50% inhibition after 30 min, activity can be restored by adding 2-mercaptoethanol
p-hydroxymercuribenzoate
phosphate
phosphoenolpyruvate
-
87% inhibition of NADP+ reduction
pyridoxal 5'-phosphate
-
5 mM, 91% inhibition of NADP+ reduction
TTP
-
2 mM, 71% inhibition of NADP+ reduction
Zn2+
; 10.1% residual activity at 2 mM
additional information
-
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2'-AMP
ADP
175.5% activity at 2 mM ADP; 75.5% activation at 2 mM
AMP
171.8% activity at 2 mM AMP; 71.8% activation at 2 mM
ATP
153.4% activity at 2 mM ATP; 53.4% activation at 2 mM
p-hydroxymercuribenzoate
-
activation, depending on presence of oxidized or reduced substrates
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.4
deamino-NAD+
-
cosubstrate NADPH
0.0003 - 0.0025
FAD
0.11 - 0.38
NAD+
0.025 - 0.085
NADH
0.015 - 0.0683
NADPH
0.04 - 0.25
thio-NAD+
0.03
thio-NADP+
-
cosubstrate NADH
additional information
additional information
-
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
167.9 - 333
NADPH
259.5
thio-NAD+
at pH 7.5 and 35°C; pH 7.5, 35°C
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2460
NADPH
at pH 7.5 and 35°C
1940
thio-NAD+
at pH 7.5 and 35°C
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0094
-
activity in inside-out mitochondrial particles from leaf
0.0131
-
activity in inside-out mitochondrial particles from tuber
24
Chromatium sp.
-
-
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
-
around pH 7.0
7
-
NADH formation
9.6
-
NADPH formation
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pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 10.5
-
pH 7.0: about 55% of maximal activity, pH 10.5: about 45% of maximal activity
7 - 9
more than 50% activity between pH 7.0 and 9.0
8.4 - 8.7
-
half maximal activity in this pH range in the absence of Ca2+ or Mg2+, no activity above
9.1 - 9.3
-
half maximal activity in this pH range in the presence of 5 mM Ca2+ or Mg2+, no activity above
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TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20 - 45
more than 70% activity between 20 and 45°C
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SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
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MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
51500
x * 51500, calculated from amino acid sequence
53000
-
SDS-PAGE
58000
-
x * 58000, SDS-PAGE
421000
1600000
-
sedimentation equilibrium in presence of 2'-AMP
6400000
-
sedimentation equilibrium in presence of NADP+
27000000
-
rod-like polymers, electron micrography
30000000 - 50000000
-
rod-like structure, light scattering
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SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
x * 51500, calculated from amino acid sequence; x * 52000, SDS-PAGE
octamer
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Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
-
polymeric enzyme stable
392577
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 50
the enzyme retains 50% activity after 5 h at 50°C. The enzyme is stable at 4°C for 25 days and retains 65% activity at 25°C. The enzyme is rapidly inactivated at high temperatures, retaining 80%, 50% and 10% activity after incubation for 30 min at 50, 57 and 62°C, respectively
51
-
25 min, 50% inactivation, accelerated by addition of NADPH, reactivation by FAD
55
-
approx. 50% activity lost after about 2 min, almost complete loss of activity after 20 min, biphasic inactivation: 70% activity lost with a first-order inactivation constant, 30% is lost much more rapidly, rate of thermal inactivation depends on concentration of NAD+, NADP+, NADH, NADPH, free FAD, Mg2+ and phosphate, independent of pH between pH 5 and pH 9, significant acceleration outside this range, addition of 1 mM FAD lowers inactivation rate about 20fold
57
purified enzyme, pH 7.5, 30 min, 50% activity remaining
62
purified enzyme, pH 7.5, 30 min, 10% activity remaining
65
-
15 min, complete inactivation, protection by FAD
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GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
50% inactivation after 10 min in 1 M guanidinium HCl
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bovine serum albumin, 0.2%, stabilization of diluted solutions
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urea, 5 min, 50% inactivation
-
urea, 8 M, no dissociation, complete inactivation of enzyme activity in 5 M urea
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-15°C, 50 mM Tris-HCl buffer, pH 7.5, no loss of activity in 12 weeks
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-20°C, 50 mM Tris-HCl, pH 7.0, 2 mM dithiothreitol, several weeks, no loss of activity, great loss of activity after several months
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25°C, purified enzyme, 25 days, 65% activity remaining
4°C, 0.1 M phosphate buffer, pH 7.5, 1 mM EDTA, several months, no loss of activity, storage at -20°C yields a partly insoluble enzyme
-
4°C, purified enzyme, 25 days, stable
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
affinity chromatography
-
affinity chromatography on immobilized 2'-AMP
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affinity chromatography, native and recombinant enzyme
-
TALON metal affinity resin column chromatography
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Escherichia coli strain is transformed with a two plasmid system, one encoding the udhA gene and the other one encoding the phb operon
-
expressed in Escherichia coli DH5alpha cells; expression of the soluble enzyme in Escherichia coli strain DH5alpha
expression in Escherichia coli
-
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
F215S
-
the mutation is associated with familial glucocorticoid deficiency
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Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
inactivation by 4 M guanidinium HCl, 4-6% reactivation after transfer in 100 mM Tris buffer, pH 7.5 containing 100 mM FAD
-
urea inactivation: 3% reactivation after dialysis against buffer containing 0.1 mM FAD, 35% after dialysis against buffer containing 1% mercaptoethanol and 95% reactivation after dialysis against buffer containing both 0.1 mM FAD and 1% mercaptoethanol
-
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
biotechnology
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Show AA Sequence (2734 entries)
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LINKS TO OTHER DATABASES (specific for EC-Number 1.6.1.1)
ExplorEnz
ExPASy
KEGG
MetaCyc
SABIO-RK
NCBI: PubMed, Protein, Nucleotide, Structure, Genome, OMIM
IUBMB Enzyme Nomenclature
PROSITE Database of protein families and domains
InterPro (database of protein families, domains and functional sites)