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Information on EC 1.8.1.4 - dihydrolipoyl dehydrogenase and Organism(s) Escherichia coli and UniProt Accession P0A9P0

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IUBMB Comments
A flavoprotein (FAD). A component of the multienzyme 2-oxo-acid dehydrogenase complexes. In the pyruvate dehydrogenase complex, it binds to the core of EC 2.3.1.12, dihydrolipoyllysine-residue acetyltransferase, and catalyses oxidation of its dihydrolipoyl groups. It plays a similar role in the oxoglutarate and 3-methyl-2-oxobutanoate dehydrogenase complexes. Another substrate is the dihydrolipoyl group in the H-protein of the glycine-cleavage system ({AminoAcid/GlyCleave} for diagram), in which it acts, together with EC 1.4.4.2, glycine dehydrogenase (decarboxylating), and EC 2.1.2.10, aminomethyltransferase, to break down glycine. It can also use free dihydrolipoate, dihydrolipoamide or dihydrolipoyllysine as substrate. This enzyme was first shown to catalyse the oxidation of NADH by methylene blue; this activity was called diaphorase. The glycine cleavage system is composed of four components that only loosely associate: the P protein (EC 1.4.4.2), the T protein (EC 2.1.2.10), the L protein (EC 1.8.1.4) and the lipoyl-bearing H protein .
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Escherichia coli
UNIPROT: P0A9P0
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The taxonomic range for the selected organisms is: Escherichia coli
The enzyme appears in selected viruses and cellular organisms
Synonyms
lipoamide dehydrogenase, dihydrolipoamide dehydrogenase, dldh, l-protein, dihydrolipoyl dehydrogenase, nadh diaphorase, e3 component, lipdh, lipoyl dehydrogenase, nicotinamide adenine dinucleotide diaphorase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dihydrolipoamide dehydrogenase
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dehydrogenase, lipoamide
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dehydrolipoate dehydrogenase
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DHLDH
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diaphorase
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dihydrolipoamide dehydrogenase
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dihydrolipoamide dehydrogenase E3
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common component of the three 2-oxoacid dehydrogenase complexes oxidizing pyruvate, 2-oxoglutarate, and the branched-chain 2-oxo acids
dihydrolipoic dehydrogenase
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dihydrolipoyl dehydrogenase
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DLDH
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-
-
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E3
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-
-
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E3 component of 2-oxoglutarate dehydrogenase complex
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-
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E3 component of acetoin cleaving system
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-
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E3 component of alpha keto acid dehydrogenase complexes
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E3 component of pyruvate and 2-oxoglutarate dehydrogenases complexes
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E3 component of pyruvate complex
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E3 lipoamide dehydrogenase
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Glycine cleavage system L protein
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Glycine oxidation system L-factor
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-
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LDP-Glc
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-
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LDP-Val
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lipoamide dehydrogenase (NADH)
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lipoamide oxidoreductase (NADH)
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lipoamide reductase
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lipoate dehydrogenase
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lipoic acid dehydrogenase
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lipoyl dehydrogenase
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LPD-GLC
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LPD-VAL
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NADH:lipoamide oxidoreductase
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-
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ORF-E3
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
protein N6-(dihydrolipoyl)lysine + NAD+ = protein N6-(lipoyl)lysine + NADH + H+
show the reaction diagram
in the physiological direction, dihydrolipoamide, which is covalently tethered to another enzymatic subunit in the multienzyme complex, binds to the disulfide-exchange site near the si face of the FAD cofactor. Dihydrolipoamide is thought to donate a hydride to the disulfide and a proton to an active-site base forming a stable charge-transfer complex between the thiolate of the mixed disulfide and the oxidized FAD cofactor. In the presence of NAD+, electrons are passed to FAD and then to NAD+ on the re face of the flavin, forming NADH with the release of a proton, mechanism modelling, detailed overview
protein N6-(dihydrolipoyl)lysine + NAD+ = protein N6-(lipoyl)lysine + NADH + H+
show the reaction diagram
ping-pong mechanism
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
redox reaction
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oxidation
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reduction
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SYSTEMATIC NAME
IUBMB Comments
protein-N6-(dihydrolipoyl)lysine:NAD+ oxidoreductase
A flavoprotein (FAD). A component of the multienzyme 2-oxo-acid dehydrogenase complexes. In the pyruvate dehydrogenase complex, it binds to the core of EC 2.3.1.12, dihydrolipoyllysine-residue acetyltransferase, and catalyses oxidation of its dihydrolipoyl groups. It plays a similar role in the oxoglutarate and 3-methyl-2-oxobutanoate dehydrogenase complexes. Another substrate is the dihydrolipoyl group in the H-protein of the glycine-cleavage system ({AminoAcid/GlyCleave} for diagram), in which it acts, together with EC 1.4.4.2, glycine dehydrogenase (decarboxylating), and EC 2.1.2.10, aminomethyltransferase, to break down glycine. It can also use free dihydrolipoate, dihydrolipoamide or dihydrolipoyllysine as substrate. This enzyme was first shown to catalyse the oxidation of NADH by methylene blue; this activity was called diaphorase. The glycine cleavage system is composed of four components that only loosely associate: the P protein (EC 1.4.4.2), the T protein (EC 2.1.2.10), the L protein (EC 1.8.1.4) and the lipoyl-bearing H protein [6].
CAS REGISTRY NUMBER
COMMENTARY hide
9001-18-7
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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
dihydrolipoamide + NAD+
lipoamide + NADH + H+
show the reaction diagram
-
-
-
r
protein N6-(lipoyl)lysine + NADH + H+
protein N6-(dihydrolipoyl)lysine + NAD+
show the reaction diagram
-
-
-
r
2 ferricyanide + NADH
2 ferrocyanide + NAD+ + H+
show the reaction diagram
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activity with wild-type enzyme and mutant enzymes C44S and C49S
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-
?
dihydrolipoamide + NAD+
lipoamide + NADH
show the reaction diagram
O2 + NADH
?
show the reaction diagram
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activity with wild-type enzyme and mutant enzymes C44S and C49S
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-
?
oxidized 2,6-dichlorophenolindophenol + NADH
? + NAD+
show the reaction diagram
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activity with wild-type enzyme and mutant enzymes C44S and C49S
-
-
?
tellurite + NADH
NAD+ + ?
show the reaction diagram
-
-
-
-
?
thio-NAD+ + NADH
thio-NADH + NAD+
show the reaction diagram
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activity with wild-type enzyme and mutant enzymes C44S and C49S
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-
?
additional information
?
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NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
dihydrolipoamide + NAD+
lipoamide + NADH + H+
show the reaction diagram
-
-
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r
protein N6-(lipoyl)lysine + NADH + H+
protein N6-(dihydrolipoyl)lysine + NAD+
show the reaction diagram
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r
additional information
?
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
FAD
flavoenzyme
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
arsenite
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in presence of NADH, inhibition is reversed by dithiols and less effectively by monothiols
Cd2+
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in presence of NADH, inhibition is reversed by dithiols and less effectively by monothiols
p-[(bromoacetyl)-amino]phenyl arsenoxide
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irreversible active site directed inactivation
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.025 - 0.28
dihydrolipoamide
0.15 - 1.83
NAD+
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
additional information
-
-
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.009 - 0.016
NADH
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.5 - 8.5
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pH 6.5: about 40% of maximal activity, pH 8.5: about 50% of maximal activity
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
UniProt
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
dihydrolipoamide dehydrogenase is a member of the disulfide oxidoreductase family
metabolism
dihydrolipoamide dehydrogenase (E3) is a component of three different catabolic multienzyme complexes that oxidize pyruvate, 2-oxoglutarate, or glycine, where E3 catalyzes the final step in a sequence of oxidative reactions
physiological function
in vivo, the dihydrolipoamide dehydrogenase component (E3) is associated with the pyruvate, 2-oxoglutarate, and glycine dehydrogenase complexes. The pyruvate dehydrogenase (PDH) complex connects the glycolytic flux to the tricarboxylic acid cycle and is central to the regulation of primary metabolism. Regulation of PDH via regulation of the E3 component by the NAD+/NADH ratio represents one of the important physiological control mechanisms of PDH activity. Steady-state distributions of enzyme redox states as a function of lipoamide/ dihydrolipoamide, NAD+/NADH, and pH, modelling, overview
physiological function
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dihydrolipoamide dehydrogenase of Escherichia coli is a bacterial enzyme that is involved in the central metabolism and shared in common between the pyruvate dehydrogenase and 2-oxoglutarate dehydrogenase complexes. The presence of oligomeric forms of the enzyme is determined by the multifunctionality of LpD in the cell, in particular, the required stoichiometry in the complexes. The E3 enzyme activity is essential for aerobic respiration. Dihydrolipoamide dehydrogenase plays an equally important role in anaerobic organisms, since this enzyme is involved in the synthesis of branched-chain keto and amino acids
additional information
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all of the E3 enzymes function as dimers, and their active site contains the reactive disulfide bridge, which is directly involved in catalysis
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
46000
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x * 46000, SDS-PAGE
51274
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x * 51274, calculation from nucleotide sequence
56000
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2 * 56000, SDS-PAGE
88000
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gel filtration
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
homodimer
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additional information
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in solution LpD exists as an equilibrium mixture of a dimer and a tetramer, small-angle X-ray scattering and analytical ultracentrifugation
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified enzyme, X-ray diffraction structure determination and analysis, small-angle X-ray scattering of the enzyme in solution, measurements in 50 mM Tris buffer, pH 7.5, at 10°C, molecular docking and modeling
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C44S
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0.003% of the activity of wild-type enzyme with NAD+ and dihydrolipoamide. Enzyme is capable to catalyze reactions with NADH as electron donor and ferricyanide, thio-NAD+, 2,6-dichlorophenol indophenol and O2 as electron acceptor. The fluorescence of FAD in oxidized wild-type enzyme is markedly temperature dependent, while the fluorescence of FAD in mutants C44S and C49S is independent of temperature
C49S
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0.012% of the activity of wild-type enzyme with NAD+ and dihydrolipoamide. Enzyme is capable to catalyze reactions with NADH as electron donor and ferricyanide, thio-NAD+, 2,6-dichlorophenol indophenol and O2 as electron acceptor. The fluorescence of FAD in oxidized wild-type enzyme is markedly temperature dependent, while the fluorescence of FAD in mutants C44S and C49S is independent of temperature
K53R
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spectral and redox properties of FAD in the mutant enzyme as well as the interaction of the flavin with bound NAD+ are profoundly affected by the mutation, K53R does not catalyze either the dihydrolipoamide-NAD+ or the NADH-lipoamide reactions except at very low concentrations of reducing substrate. The absorbance spectrum in the visible and near-ultraviolet is little changed from that of wild-type enzyme, in contrast to wild-type enzyme the spectrum of K53R is sensitive to pH. Unlike the wild-type enzyme, the binding of beta-NAD+ to K53R alters the spectrum
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
from pyruvate dehydrogenase complex
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purified from strain ASKA clone (-) JW0112
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recombinant enzyme from Escherichia coli strain BL21(DE3) by ammonium sulfate fractionation and hydrophobic interaction chromatography to over 98% purity
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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21(DE3)
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gene lpd, recombinant expression in Escherichia coli strain BL21(DE3)
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
degradation
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shows NADH-dependent tellurite reductase activity in vitro
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Reed, L.J.; Willms, C.R.
Purification and resolution of the pyruvate dehydrogenase complex (Escherichia coli)
Methods Enzymol.
9
247-265
1966
Escherichia coli
-
Manually annotated by BRENDA team
Richarme, G.
Purification of a new dihydrolipoamide dehydrogenase from Escherichia coli
J. Bacteriol.
171
6580-6585
1989
Escherichia coli
Manually annotated by BRENDA team
Carothers, D.J.; Pons, G.; Patel, M.S.
Dihydrolipoamide dehydrogenase: functional similarities and divergent evolution of the pyridine nucleotide-disulfide oxidoreductases
Arch. Biochem. Biophys.
268
409-425
1989
Ascaris suum, Azotobacter vinelandii, Bacillus subtilis, Bos taurus, Escherichia coli, Geobacillus stearothermophilus, Halobacterium salinarum, Homo sapiens, Pisum sativum, Pseudomonas aeruginosa, Pseudomonas putida, Rattus norvegicus, Saccharomyces cerevisiae, Saccharomyces pastorianus, Sus scrofa
Manually annotated by BRENDA team
Adamson, S.R.; Stevenson, K.J.
Inhibition of pyruvate dehydrogenase multienzyme complex from Escherichia coli with a bifunctional arsenoxide: selective inactivation of lipoamide dehydrogenase
Biochemistry
20
3420-3424
1981
Escherichia coli
-
Manually annotated by BRENDA team
Schmincke-Ott, E.; Bisswanger, H.
Dihydrolipoamide dehydrogenase component of the pyruvate dehydrogenase complex from Escherichia coli K12. Comparative characterization of the free and the complex-bound component
Eur. J. Biochem.
114
413-420
1981
Escherichia coli
Manually annotated by BRENDA team
Hopkins, N.; Williams, C.H., Jr.
Characterization of lipoamide dehydrogenase from Escherichia coli lacking the redox active disulfide: C44S and C49S
Biochemistry
34
11757-11765
1995
Escherichia coli
Manually annotated by BRENDA team
Maeda-Yorita, K.; Russell, G.C.; Guest, J.R.; Massey, V.; Williams, C.H., Jr.
Modulation of the Oxidation-Reduction Potential of the Flavin in Lipoamide Dehydrogenase from Escherichia coli by Alteration of a Nearby Charged Residue, K53R
Biochemistry
33
6213-6220
1994
Escherichia coli
Manually annotated by BRENDA team
Williams, C.H.
Flavin-containing dehydrogenases
The Enzymes, 3rd Ed. (Boyer, P. D. , ed. )
13
89-173
1976
Azotobacter agilis, Azotobacter vinelandii, Bacillus subtilis, Bos taurus, Brassica oleracea, Enterococcus faecalis, Escherichia coli, Escherichia coli B / ATCC 11303, Escherichia coli Crookes, Escherichia coli M191-6, Globisporangium ultimum, Homo sapiens, Leuconostoc mesenteroides, Mycobacterium tuberculosis, Neurospora crassa, Parvimonas micra, Phytophthora erythroseptica, Pichia kudriavzevii, Proteus vulgaris, Pseudomonas fluorescens, Rattus norvegicus, Saccharomyces cerevisiae, Serratia marcescens, Spinacia oleracea, Squalus acanthias, Sus scrofa
-
Manually annotated by BRENDA team
Conner, M.; Krell, T.; Lindsay, J.G.
Identification and purification of a distinct dihydrolipoamide dehydrogenase from pea chloroplasts
Planta
200
195-202
1996
Escherichia coli, Pisum sativum
Manually annotated by BRENDA team
Scouten, W.H.; McManus, I.R.
Microbial lipoamide dehydrogenase. Purification and some characteristics of the enzyme derived from selected microorganisms
Biochim. Biophys. Acta
227
248-263
1971
Azotobacter agilis, Bacillus subtilis, Saccharomyces cerevisiae, Escherichia coli, Neurospora crassa, Pseudomonas fluorescens, Serratia marcescens
Manually annotated by BRENDA team
Castro, M.E.; Molina, R.; Diaz, W.; Pichuantes, S.E.; Vasquez, C.C.
The dihydrolipoamide dehydrogenase of Aeromonas caviae ST exhibits NADH-dependent tellurite reductase activity
Biochem. Biophys. Res. Commun.
375
91-94
2008
Aeromonas caviae, Escherichia coli, Geobacillus stearothermophilus (P11959), Geobacillus stearothermophilus, Zymomonas mobilis (P50970), Zymomonas mobilis, Streptococcus pneumoniae (Q8VPK7), Streptococcus pneumoniae
Manually annotated by BRENDA team
Moxley, M.A.; Beard, D.A.; Bazil, J.N.
A pH-dependent kinetic model of dihydrolipoamide dehydrogenase from multiple organisms
Biophys. J.
107
2993-3007
2014
Spinacia oleracea (A0A0K9R8G5), Spinacia oleracea, Homo sapiens (P09622), Homo sapiens, Escherichia coli (P0A9P0), Escherichia coli
Manually annotated by BRENDA team
Dadinova, L.; Rodina, E.; Vorobyeva, N.; Kurilova, S.; Nazarova, T.; Shtykova, E.
Structural investigations of E. coli dihydrolipoamide dehydrogenase in solution small-angle X-ray scattering and molecular docking
Crystallogr. Rep.
61
414-420
2016
Escherichia coli
-
Manually annotated by BRENDA team