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Information on EC 2.3.1.12 - dihydrolipoyllysine-residue acetyltransferase and Organism(s) Escherichia coli and UniProt Accession P06959

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IUBMB Comments
A multimer (24-mer or 60-mer, depending on the source) of this enzyme forms the core of the pyruvate dehydrogenase multienzyme complex, and binds tightly both EC 1.2.4.1, pyruvate dehydrogenase (acetyl-transferring) and EC 1.8.1.4, dihydrolipoyl dehydrogenase. The lipoyl group of this enzyme is reductively acetylated by EC 1.2.4.1, and the only observed direction catalysed by EC 2.3.1.12 is that where the acetyl group is passed to coenzyme A.
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Escherichia coli
UNIPROT: P06959
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The taxonomic range for the selected organisms is: Escherichia coli
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota, Archaea
Synonyms
dihydrolipoamide acetyltransferase, dihydrolipoyl transacetylase, lipoate acetyltransferase, dihydrolipoyl acetyltransferase, dhlta, dihydrolipoyl acetyl transferase, dihydrolipoyllysine-residue acetyltransferase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dihydrolipoamide acetyltransferase
-
dihydrolipoyl acetyltransferase
-
pyruvate dehydrogenase complex component E2
-
pyruvate dehydrogenase complex component E2p
-
acetyltransferase, lipoate
-
-
-
-
DHLTA
-
-
-
-
dihydrolipoamide acetyltransferase
-
-
dihydrolipoate acetyltransferase
-
-
-
-
dihydrolipoic transacetylase
-
-
-
-
dihydrolipoyl acetyltransferase
-
-
-
-
E2p
-
-
-
-
lipoate acetyltransferase
-
-
-
-
lipoate transacetylase
-
-
-
-
lipoic acetyltransferase
-
-
-
-
lipoic acid acetyltransferase
-
-
-
-
lipoic transacetylase
-
-
-
-
lipoylacetyltransferase
-
-
-
-
myelin-proteolipid O-palmitoyltransferase
-
-
-
-
palmitoyl-CoA:myelin-proteolipid O-palmitoyltransferase
-
-
-
-
thioltransacetylase A
-
-
-
-
transacetylase X
-
-
-
-
additional information
-
the enzyme is a subunit of the pyruvate dehydrogenase multienzyme complex
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acyl group transfer
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
acetyl-CoA:enzyme N6-(dihydrolipoyl)lysine S-acetyltransferase
A multimer (24-mer or 60-mer, depending on the source) of this enzyme forms the core of the pyruvate dehydrogenase multienzyme complex, and binds tightly both EC 1.2.4.1, pyruvate dehydrogenase (acetyl-transferring) and EC 1.8.1.4, dihydrolipoyl dehydrogenase. The lipoyl group of this enzyme is reductively acetylated by EC 1.2.4.1, and the only observed direction catalysed by EC 2.3.1.12 is that where the acetyl group is passed to coenzyme A.
CAS REGISTRY NUMBER
COMMENTARY hide
9032-29-5
-
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
acetyl-CoA + enzyme N6-(dihydrolipoyl)lysine
CoA + enzyme N6-(S-acetyldihydrolipoyl)lysine
show the reaction diagram
CoA + enzyme N6-(S-acetyldihydrolipoyl)lysine
acetyl-CoA + enzyme N6-(dihydrolipoyl)lysine
show the reaction diagram
-
-
-
r
acetyl-CoA + enzyme N6-(dihydrolipoyl)lysine
CoA + enzyme N6-(S-acetyldihydrolipoyl)lysine
show the reaction diagram
-
reductive acetylation of the enzyme
-
-
?
dihydrolipoamide + acetyl-CoA
S-acetyldihydrolipoamide + CoA
show the reaction diagram
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
acetyl-CoA + enzyme N6-(dihydrolipoyl)lysine
CoA + enzyme N6-(S-acetyldihydrolipoyl)lysine
show the reaction diagram
dihydrolipoamide + acetyl-CoA
S-acetyldihydrolipoamide + CoA
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
thiamine diphosphate
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Iodine
-
complete inactivation at 2.5 mol iodine per mol of subunit
N-Acetylimidazole
-
modifies tyrosyl residues and dissociates the enzyme into subunits
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
4.2
dihydrolipoamide
-
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
34.1
-
purified 1-lip E2
47.1
-
catalytic domain
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8
-
assay at
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25
-
assay at
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30028
x * 30028, catalytic domain, calculated from amino acid sequence
30030
x * 30030, catalytic domain, FT mass spectrometry
65959
x * 65959, calculated from amino acid sequence
1000000
-
sedimentation equilibrium centrifugation
1470000
-
sedimentation equilibrium experiments in 0.83 M acetic and and 0.005 M sodium chloride
150000
-
no component of pyruvate dehydrogenase complex, sucrose density gradient centrifugation
1548000
-
native enzyme
36000
40000
-
24 * 40000, linked by noncovalent bonds
45953
-
x * 45953, recombinant 1-lip E2, mass spectrometry, x * 8982, recombinant unacetylated hybrid lipoyl domain, mass spectrometry, x * 9019, recombinant fully acetylated hybrid lipoyl domain, mass spectrometry
60100 - 64500
-
sedimentation equilibrium, depending on buffer
65959
-
24 * 65959, calculation from nucleotide sequence
65960
-
calculation from nucleotide sequence
670000
-
gel filtration
78000
-
SDS-PAGE
80000
81000
-
SDS-PAGE, gel filtration
83000
-
wild-type enzyme, SDS-PAGE
864000
-
crystallization experiments
8982
-
x * 45953, recombinant 1-lip E2, mass spectrometry, x * 8982, recombinant unacetylated hybrid lipoyl domain, mass spectrometry, x * 9019, recombinant fully acetylated hybrid lipoyl domain, mass spectrometry
9019
-
x * 45953, recombinant 1-lip E2, mass spectrometry, x * 8982, recombinant unacetylated hybrid lipoyl domain, mass spectrometry, x * 9019, recombinant fully acetylated hybrid lipoyl domain, mass spectrometry
additional information
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
trimer
x-ray crystallography
?
-
x * 45953, recombinant 1-lip E2, mass spectrometry, x * 8982, recombinant unacetylated hybrid lipoyl domain, mass spectrometry, x * 9019, recombinant fully acetylated hybrid lipoyl domain, mass spectrometry
dimer
polymer
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
lipoprotein
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
E1p-E2p didomain subcomplex, sitting drop vapor diffusion method, using 20% (w/v) PEG 3350, 0.2% (w/v) NaN3, and 0.2 M ammonium tartrate dibasic buffer (pH 6.35)
His9-tagged catalytic domain, hanging drop vapor diffusion method, using 0.2 M potassium thiocyanate and 20% (w/v) polyethylene glycol 3350
E2-E3 subcomplex of pyruvate dehydrogenase, E2: EC 2.3.1.12, E3: EC 1.8.1.4, various crystallization conditions
-
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
H602C
inactive
K191E
the mutant shows 50% activity compared to the wild type enzyme
K191R
the mutant shows 79% activity compared to the wild type enzyme
R129A
the mutant shows 34% activity compared to the wild type enzyme
R129E
the mutant shows 3.0% activity compared to the wild type enzyme
R129K
the mutant shows 77% activity compared to the wild type enzyme
R130A
the mutant shows wild type activity
R130E
the mutant shows 57% activity compared to the wild type enzyme
R150A
the mutant shows 8.0% activity compared to the wild type enzyme
R150E
the mutant shows 2.4% activity compared to the wild type enzyme
R150K
the mutant shows 5.0% activity compared to the wild type enzyme
R153E
the mutant shows 60% activity compared to the wild type enzyme
R153K
the mutant shows 83% activity compared to the wild type enzyme
R161A
the mutant shows 65% activity compared to the wild type enzyme
R161E
the mutant shows 84% activity compared to the wild type enzyme
R202E
the mutant shows 10% activity compared to the wild type enzyme
R202K
the mutant shows 86% activity compared to the wild type enzyme
K244Q
-
unlipoylatable domain
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3 - 11
-
isolated E. coli lipoyl domains stay soluble in this range
486166
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
70
-
1 h stable, isolated lipoyl domains
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
nickel affinity column chromatography
nickel affinity resin column chromatography, Superdex G75 gel filtration, and G3000SW TSK gel filtration
E1-E2 complex
-
fusion protein of 6 amino acids form beta-galactosidase, the apa-4 region and the catalytic domain of E2
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isolation of lipoyl domains
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pyruvate dehydrogenase complex and tryptic fragments of E2
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pyruvate dehydrogenase complex, stoichiometry E1:E2:E3 is 1.56:1:0.89
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recombinant 1-lip E2 and recombinant hybrid lipoyl domain by ammonium sulfate fractionation, ion exchange and hydrophobic interaction chromatography
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wild-type and mutants with deletions of lipolyl domains
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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli AG1 cells
expressed in Escherichia coli BL21(DE3) cells
functional overexpression of the enzyme E2 with a single hybrid lipoyl domain per subunit, i.e. 1-lip E2, in strain JRG1342 and BL21(DE3), overexpression of the isolated hybrid lipoyl domain in strain JM 101
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wild-type and mutants
-
RENATURED/Commentary
ORGANISM
UNIPROT
LITERATURE
reconstitution of E1 with E2-E3 complex with a stoichiometry of E1:E2:E3 of 2:1:1
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reconstitution of pyruvate dehydrogenase complex, maximum activity is produced when transacetylase accommodates 12 pyruvate dehydrogenase dimers and 6 flavoprotein dimers
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reconstitution of pyruvate dehydrogenase complex, stoichiometry pyruvate dehydrogenase:transacetylase:flavoprotein is 1:0.35:0.4
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reconstitution of the active multienzyme complex with recombinant components
-
treatment with dilute acetic acid solution results in dissociation into inactive subunits with MW 70000, removal of the acid results in restoration of enzymatic activity
-
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
De Kok, A.; Westphal, A.H.
Hybrid pyruvate dehydrogenase complexes reconstituted from components of the complexes from Escherichia coli and Azotobacter vinelandii
Eur. J. Biochem.
152
35-41
1985
Azotobacter vinelandii, Escherichia coli
Manually annotated by BRENDA team
Graupe, K.; Abusaud, M.; Karfunkel, H.; Bisswanger, H.
Reassociation of the pyruvate dehydrogenase complex from Escherichia coli: Kinetic measurements and binding studies by resonance energy transfer
Biochemistry
21
1386-1394
1982
Escherichia coli
Manually annotated by BRENDA team
Guest, J.R.; Stephens, P.E.
Molecular cloning of the pyruvate dehydrogenase complex genes of Escherichia coli
J. Gen. Microbiol.
121
277-292
1980
Escherichia coli
Manually annotated by BRENDA team
Danson, M.J.; Hale, G.; Johnson, P.; Perham, R.N.
Molecular weight and symmetry of the pyruvate dehydrogenase multienzyme complex of Escherichia coli
J. Mol. Biol.
129
603-617
1979
Escherichia coli
Manually annotated by BRENDA team
Guest, J.R.; Darlison, M.G.; Spencer, M.E.; Stephens, P.E.
Cloning and sequence analysis of the pyruvate and 2-oxoglutarate dehydrogenase complex genes of Escherichia coli
Biochem. Soc. Trans.
12
220-223
1984
Escherichia coli
Manually annotated by BRENDA team
Ali, S.T.; Guest, J.R.
Isolation and characterization of lipoylated and unlipoylated domains of the E2p subunit of the pyruvate dehydrogenase complex of Escherichia coli
Biochem. J.
271
139-145
1990
Escherichia coli
Manually annotated by BRENDA team
Guest, J.R.; Lewis, H.M.; Graham, L.D.; Packman, L.C.; Perham, R.N.
Genetic reconstruction and functional analysis of the repeating lipoyl domains in the pyruvate dehydrogenase multienzyme complex of Escherichia coli
J. Mol. Biol.
185
743-754
1985
Escherichia coli
Manually annotated by BRENDA team
Packman, L.C.; Hale, G.; Perham, R.N.
Repeating functional domains in the pyruvate dehydrogenase multienzyme complex of Escherichia coli
EMBO J.
3
1315-1319
1984
Escherichia coli
Manually annotated by BRENDA team
Danson, M.J.; Porteous, C.E.
Pyruvate dehydrogenase multienzyme complex of Escherichia coli: determination of the Mr of the lipoate acetyltransferase component
FEBS Lett.
133
112-114
1981
Escherichia coli
Manually annotated by BRENDA team
Bleile, D.M.; Munk, P.; Oliver, R.M.; Reed, L.J.
Subunit structure of dihydrolipoyl transacetylase component of pyruvate dehydrogenase complex from Escherichia coli
Proc. Natl. Acad. Sci. USA
76
4385-4389
1979
Escherichia coli
Manually annotated by BRENDA team
Fuller, C.C.; Reed, L.J.; Oliver, R.M.; Hackert, M.L.
Crystallization of a dihydrolipoyl transacetylase - dihydrolipoyl dehydrogenase subcomplex and its implications regarding the subunit structure of the pyruvate dehydrogenase complex from Escherichia coli
Biochem. Biophys. Res. Commun.
90
431-438
1979
Escherichia coli
Manually annotated by BRENDA team
Reed, L.J.; Pettit, F.H.; Eley, M.H.; Hamilton, L.; Collins, J.H.; Oliver, R.M.
Reconstitution of the Escherichia coli pyruvate dehydrogenase complex
Proc. Natl. Acad. Sci. USA
72
3068-3072
1975
Escherichia coli
Manually annotated by BRENDA team
Bisswanger, H.; Henning, U.
A new dihydrolipoamide transacetylase in Escherichia coli K12
Biochim. Biophys. Acta
321
143-148
1973
Escherichia coli
Manually annotated by BRENDA team
Vogel, O.; Beikirch, H.; Muller, H.; Henning, U.
The subunit structure of the Escherichia coli K-12 pyruvate dehydrogenase complex. The dihydrolipoamide transacetylase component
Eur. J. Biochem.
20
169-178
1971
Escherichia coli
Manually annotated by BRENDA team
Schwartz, E.R.; Reed, L.J.
alpha-Keto acid dehydrogenase complexes. XII. Effects of acetylation on the activity and structure of the dihydrolipoyl transacetylase of Escherichia coli
J. Biol. Chem.
244
6074-6079
1969
Escherichia coli
Manually annotated by BRENDA team
Schwartz, E.; Reed, L.J.
alpha-Keto acid dehydrogenase complexes. IX. Effects of iodination of the tyrosyl residues on the properties of the dihydrolipoyl transacetylase of Escherichia coli
J. Biol. Chem.
243
639-643
1968
Escherichia coli
Manually annotated by BRENDA team
Henney, H.R.; Willms, C.R.; Muramatsu, T.; Mukherjee, B.B.; Reed, L.J.
alpha-Keto acid dehydrogenase complexes. VII. Isolation and partial characterization of the polypeptide chains in the dihydrolipoyl transacetylase of Escherichia coli
J. Biol. Chem.
242
898-901
1967
Escherichia coli
Manually annotated by BRENDA team
Willms, C.R.; Oliver, R.M.; Henney, H.R.; Mukherjee, B.B.; Reed, L.J.
alpha-Keto acid dehydrogenase complexes. VI. Dissociation and reconstitution of the dihydrolipoyl transacetylase of Escherichia coli
J. Biol. Chem.
242
889-897
1967
Escherichia coli
Manually annotated by BRENDA team
Schulze, E.; Westphal, A.H.; Obmolova, G.; Mattevi, A.; Hol, W.G.; de Kok, A.
The catalytic domain of the dihydrolipoyl transacetylase component of the pyruvate dehydrogenase complex from Azotobacter vinelandii and Escherichia coli. Expression, purification, properties and preliminary X-ray analysis
Eur. J. Biochem.
201
561-568
1991
Azotobacter vinelandii, Escherichia coli
Manually annotated by BRENDA team
Schulze, E.; Westphal, A.H.; Veenhuis, M.; de Kok, A.
Purification and cellular localization of wild type and mutated dihydrolipoyltransacetylases from Azotobacter vinelandii and Escherichia coli expressed in E. coli
Biochim. Biophys. Acta
1120
87-96
1992
Azotobacter vinelandii, Escherichia coli
Manually annotated by BRENDA team
Wei, W.; Li, H.; Nemeria, N.; Jordan, F.
Expression and purification of the dihydrolipoamide acetyltransferase and dihydrolipoamide dehydrogenase subunits of the Escherichia coli pyruvate dehydrogenase multienzyme complex: a mass spectrometric assay for reductive acetylation of dihydrolipoamide acetyltransferase
Protein Expr. Purif.
28
140-150
2003
Escherichia coli
Manually annotated by BRENDA team
Chandrasekhar, K.; Wang, J.; Arjunan, P.; Sax, M.; Park, Y.H.; Nemeria, N.S.; Kumaran, S.; Song, J.; Jordan, F.; Furey, W.
Insight to the interaction of the dihydrolipoamide acetyltransferase (E2) core with the peripheral components in the Escherichia coli pyruvate dehydrogenase complex via multifaceted structural approaches
J. Biol. Chem.
288
15402-15417
2013
Escherichia coli (P06959), Escherichia coli
Manually annotated by BRENDA team
Wang, J.; Nemeria, N.S.; Chandrasekhar, K.; Kumaran, S.; Arjunan, P.; Reynolds, S.; Calero, G.; Brukh, R.; Kakalis, L.; Furey, W.; Jordan, F.
Structure and function of the catalytic domain of the dihydrolipoyl acetyltransferase component in Escherichia coli pyruvate dehydrogenase complex
J. Biol. Chem.
289
15215-15230
2014
Escherichia coli (P06959), Escherichia coli
Manually annotated by BRENDA team
Arjunan, P.; Wang, J.; Nemeria, N.S.; Reynolds, S.; Brown, I.; Chandrasekhar, K.; Calero, G.; Jordan, F.; Furey, W.
Novel binding motif and new flexibility revealed by structural analyses of a pyruvate dehydrogenase-dihydrolipoyl acetyltransferase subcomplex from the Escherichia coli pyruvate dehydrogenase multienzyme complex
J. Biol. Chem.
289
30161-30176
2014
Escherichia coli (P06959)
Manually annotated by BRENDA team