Information on EC 2.7.7.63 - lipoate-protein ligase

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

EC NUMBER
COMMENTARY
2.7.7.63
-
RECOMMENDED NAME
GeneOntology No.
lipoate-protein ligase
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
ATP + lipoate = diphosphate + lipoyl-AMP
show the reaction diagram
-
-
-
-
lipoyl-AMP + apoprotein = protein N6-(lipoyl)lysine + AMP
show the reaction diagram
-
-
-
-
lipoyl-AMP + apoprotein = protein N6-(lipoyl)lysine + AMP
show the reaction diagram
LplA structures are determined: LplA/lipoyl-AMP binary complex and LplA/octyl-5'-AMP/apoH-protein ternary complex. These structures represent a post-lipoate adenylation stage and a pre-lipoate transfer stage, respectively. Three large scale conformational changes in the Escherichia coli LplA structure upon completion of the lipoate adenylation reaction are found, which enable LplA to accommodate apoprotein for the second reaction
-
PATHWAY
KEGG Link
MetaCyc Link
lipoate salvage I
-
lipoate salvage II
-
lipoate--GcvH protein ligase
-
Lipoic acid metabolism
-
Metabolic pathways
-
SYSTEMATIC NAME
IUBMB Comments
ATP:lipoate adenylyltransferase
Requires Mg2+. Both 6S- and 6R-lipoates can act as substrates but there is a preference for the naturally occurring R-form. Selenolipoate, i.e. 5-(1,2-diselenolan-3-yl)pentanoic acid, and 6-sulfanyloctanoate can also act as substrates, but more slowly [2]. This enzyme is responsible for lipoylation in the presence of exogenous lipoic acid [7]. Lipoylation is essential for the function of several key enzymes involved in oxidative metabolism, including pyruvate dehydrogenase (E2 domain), 2-oxoglutarate dehydrogenase (E2 domain), the branched-chain 2-oxoacid dehydrogenases and the glycine cleavage system (H protein) [6]. This enzyme attaches lipoic acid to the lipoyl domains of these proteins, converting apoproteins into holoproteins. It is likely that an alternative pathway, involving EC 2.3.1.181, lipoyl(octanoyl) transferase and EC 2.8.1.8, lipoyl synthase, is the normal route for lipoylation [7].
SYNONYMS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
lipoate ligase
-
-
lipoate ligase
-
-
lipoate ligase like protein
-
-
lipoate-protein ligase
-
-
lipoate-protein ligase A
-
-
lipoate-protein ligase A
-
-
lipoate-protein ligase A
-
-
lipoic acid ligase
-
-
lipoic acid ligase
-
-
lipoic acid protein ligase
-
-
lipoic acid protein ligase A1
-
-
lipoyl ligase
-
-
lipoyl-protein ligase A
-
-
LPLA
P32099
-
LplA-LplB complex
-
-
LplA1
-
functional redundant with LplA2 in medium containing free lipoic acid
Lpla2
-
functional redundant with LplA1 in medium containing free lipoic acid
Oryza s. lipoate-protein ligase A
-
-
OsLPLA
-
on chromosome 8 of Oryza sativa
CAS REGISTRY NUMBER
COMMENTARY
139639-26-2
-
ORGANISM
COMMENTARY
LITERATURE
SEQUENCE CODE
SEQUENCE DB
SOURCE
; isoenzymes LipB, LplA
-
-
Manually annotated by BRENDA team
isoenzymes LPL-A and LPL-B
-
-
Manually annotated by BRENDA team
isoenzymes LplA and LipB
-
-
Manually annotated by BRENDA team
strains MG1655 and W3110; wild type and LipB knockout strains with spontaneous mutations in lplA gene
-
-
Manually annotated by BRENDA team
Japonica type
-
-
Manually annotated by BRENDA team
enzyme homologues LipL1 and LipL2
-
-
Manually annotated by BRENDA team
lipoylation occurs in vivo in Thermoplasma acidophilum but requires two proteins LplA and CTD
Swissprot
Manually annotated by BRENDA team
lipoylation occurs in vivo in Thermoplasma acidophilum but requires two proteins LplA and CTD
UniProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
malfunction
-
LplA1 knockout cannot be achieved because of the essential function of the gene/protein in asexual parasite growth
malfunction
-
LplA1Ct is able to complement an Escherichia coli LplA knockout mutant at an efficiency of 94%
metabolism
-
The LipB octanoyltransferase catalyzes the first step of lipoic acid synthesis in Escherichia coli, transfer of the octanoyl moiety from octanoyl-acyl carrier protein to the lipoyl domains of the E2 subunits of the 2-oxoacid dehydrogenases of aerobic metabolism.
metabolism
-
LplA1 plays a critical role for asexual parasite growth in vitro and in vivo.
metabolism
-
LplA1 plays a pivotal role during the development of the erythrocytic stages of the malaria parasite.
physiological function
-
LplA1 plays a pivotal role during the development of the erythrocytic stages (asexual parasite growth) of Plasmodium falciparum
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + (R)-lipoic acid + Escherichia coli apoH-protein
diphosphate + AMP + Escherichia coli (R)-lipoyl-apoH-protein
show the reaction diagram
-
-
-
-
?
ATP + 6-thio-octanoic acid
diphosphate + 6-thio-octanoyl-AMP
show the reaction diagram
-
at 326% of the rate with DL-lipoic acid
-
-
?
ATP + 8-methyl-lipoic acid
diphosphate + 8-methyl-lipoyl-AMP
show the reaction diagram
-
at 73% of the rate with DL-lipoic acid
-
-
-
ATP + D-lipoic acid
diphosphate + D-lipoyl-AMP
show the reaction diagram
-
-
-
-
?
ATP + D-lipoic acid
diphosphate + D-lipoyl-AMP
show the reaction diagram
-
-
-
-
?
ATP + D-lipoic acid
diphosphate + D-lipoyl-AMP
show the reaction diagram
-
-
-
-
?
ATP + D-lipoic acid
diphosphate + D-lipoyl-AMP
show the reaction diagram
-
at 83% of the rate with DL-lipoic acid
-
-
-
ATP + dihydro-DL-lipoic acid
diphosphate + dihydro-DL-lipoyl-AMP
show the reaction diagram
-
at 80% of the rate with DL-lipoic acid
-
-
-
ATP + DL-lipoic acid
diphosphate + DL-lipoyl-AMP
show the reaction diagram
-
-
-
-
?
ATP + DL-lipoic acid + protein
AMP + DL-lipoyl-protein
show the reaction diagram
-, P32099
-
-
-
?
ATP + DL-lipoic acid + protein
diphosphate + AMP + DL-lipoyl-protein
show the reaction diagram
-
-
-
-
?
ATP + L-lipoic acid
diphosphate + L-lipoyl-AMP
show the reaction diagram
-
at 36% of the rate with DL-lipoic acid
-
-
-
ATP + lipoate + apoprotein
AMP + diphosphate + protein N6-(lipoyl)lysine
show the reaction diagram
-
protein lipoylation, salvage pathway
-
-
?
ATP + lipoate + biotin
AMP + diphosphate + ?
show the reaction diagram
-
-
-
-
?
ATP + lipoate + H-protein
AMP + diphosphate + ?
show the reaction diagram
-
-
-
-
?
ATP + lipoate + LplA acceptor peptide 1
?
show the reaction diagram
-
-
-
-
?
ATP + lipoate + LplA acceptor peptide 2
?
show the reaction diagram
-
-
-
-
?
ATP + octanoate + pyruvate dehydrogenase subunit E2
diphosphate + AMP + octanoyl-pyruvate dehydrogenase subunit E2
show the reaction diagram
-
lipoate-protein ligase attaches octanoate to the dehydrogenase subunit and sulfur insertion protein LipA, then converts octanoate to lipoate. LipA acts on both octanoate and octanoyl-proteins
-
-
?
ATP + octanoic acid
diphosphate + octanoyl-AMP
show the reaction diagram
-
-
-
-
?
ATP + octanoic acid
diphosphate + octanoyl-AMP
show the reaction diagram
-
at 13% of the rate with DL-lipoic acid
-
-
-
ATP + selenolipoic acid
diphosphate + selenolipoyl-AMP
show the reaction diagram
-
at 12% of the rate with DL-lipoic acid
-
-
?
DL-lipoyladenylate + protein
adenylate + DL-lipoyl-protein
show the reaction diagram
-
-
-
-
?
lipoic acid + ATP + apoprotein
diphosphate + AMP + N6-(lipoyl)-lysine
show the reaction diagram
-
two-step reaction of protein lipoylation at lysine residues with lipoyl-AMP intermediate
-
-
?
lipoic acid + ATP + apoprotein
diphosphate + AMP + N6-(lipoyl)-lysine
show the reaction diagram
-
complementation assay in LplA-deficient Escherichia coli strain TM137 (2 days, 37C) using exogenous lipoate
detection of lipoylated proteins in Western blot
-
?
octanoic acid + ATP
diphosphate + octanoyl-AMP
show the reaction diagram
-
-
-
-
?
octanoyl adenylate + protein
adenylate + octanoyl-protein
show the reaction diagram
-
-
-
-
?
octanoyl-ACP + lipoyl protein
octanoylated lipoyl protein + ACP
show the reaction diagram
-
-
-
-
?
lipoic acid + ATP + apoprotein
diphosphate + AMP + N6-(lipoyl)-lysine
show the reaction diagram
-
lipoic acid is (R)-5-(1,2-dithiolan-3-yl)pentanoic acid, also called 6,8-dithiooctanoic acid or thioctic acid
-
-
?
additional information
?
-
-
LipB protein utilizes lipoyl groups generated via endogenous, LipA mediated biosynthesis and causes the accumulation of aberrantly modified octanoyl-proteins in lipoate-deficient cells
-
-
-
additional information
?
-
-
LplA potein attaches octanoate to the dehydrogenase and LipA protein then converts the octanoate to lipoate
-
-
-
additional information
?
-
-
lipoate scavenging by enzymes drives mitochondrial lipoylation, while apicoplast lipoylation relies on biosynthesis, no substrate: pyruvate dehydrogenase subunit E2
-
-
-
additional information
?
-
-
utilization of host-derived lipoyl peptides which is prerequisite for bacterial fitness and intracellular proliferation, LplA1, lipoamide as lipoate source does not dictate LplA1-supported bacterial growth, LplA1, lipoate ligase activity since overexpression rescued growth of LplA1-deficient Escherichia coli strain, LplA1, lipoylated tripeptide released from proteinase K-digested pyruvate dehydrogenase or synthetic lipoylated tripeptide (aspartate-(lipoyl-)lysine-alanine, DK(L)A, 1 microgram) serves as lipoate substrate, LplA2, no utilization of host-derived lipoyl peptides as lipoate source, LplA2, utilization of lipoamide or free lipoate as lipoate source
-
-
-
additional information
?
-
Q9HKT1, Q9HKT2
it is shown that E2 lipoyl domain peptide, LplA and CTD are physically associated. LplA and CTD exist in a ratio 1:1
-
-
-
additional information
?
-
-
LplA has no detectable ligase activity in vitro in the absence of LplB
-
-
-
additional information
?
-
-
LplA's natural protein substrates have a conserved beta-hairpin structure
-
-
-
additional information
?
-
Q9HKT1, Q9HKT2
when the Thermoplasma acidophilum E2 lipoyl domain peptide is incubated with either HisLplA or HisCTD in the presence of ATP, MgCl2 and lipoic acid, no lipoylation is detected. Incubation with both HisLplA and HisCTD in the one reaction results in the lipoylation of the E2 lipoyl domain peptide as indicated by nondenaturing PAGE
-
-
-
additional information
?
-
-
recombinant LplA1Ct is able to lipoylate apo-PDH and 2-OGDH-E2 subunits purified from Escherichia coli ATM967 as detected by Western blotting with anti-lipoic acid antibody. LplA1Ct is also able to lipoylate the recombinant chlamydial BCKDH-E2 subunit
-
-
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
ATP + lipoate + apoprotein
AMP + diphosphate + protein N6-(lipoyl)lysine
show the reaction diagram
-
protein lipoylation, salvage pathway
-
-
?
ATP + lipoate + H-protein
AMP + diphosphate + ?
show the reaction diagram
-
-
-
-
?
lipoic acid + ATP + apoprotein
diphosphate + AMP + N6-(lipoyl)-lysine
show the reaction diagram
-
two-step reaction of protein lipoylation at lysine residues with lipoyl-AMP intermediate
-
-
?
lipoic acid + ATP + apoprotein
diphosphate + AMP + N6-(lipoyl)-lysine
show the reaction diagram
-
lipoic acid is (R)-5-(1,2-dithiolan-3-yl)pentanoic acid, also called 6,8-dithiooctanoic acid or thioctic acid
-
-
?
additional information
?
-
-
LipB protein utilizes lipoyl groups generated via endogenous, LipA mediated biosynthesis and causes the accumulation of aberrantly modified octanoyl-proteins in lipoate-deficient cells
-
-
-
additional information
?
-
-
LplA potein attaches octanoate to the dehydrogenase and LipA protein then converts the octanoate to lipoate
-
-
-
additional information
?
-
-
lipoate scavenging by enzymes drives mitochondrial lipoylation, while apicoplast lipoylation relies on biosynthesis
-
-
-
additional information
?
-
-
utilization of host-derived lipoyl peptides which is prerequisite for bacterial fitness and intracellular proliferation
-
-
-
additional information
?
-
Q9HKT1, Q9HKT2
it is shown that E2 lipoyl domain peptide, LplA and CTD are physically associated. LplA and CTD exist in a ratio 1:1
-
-
-
additional information
?
-
-
LplA has no detectable ligase activity in vitro in the absence of LplB
-
-
-
additional information
?
-
-
LplA's natural protein substrates have a conserved beta-hairpin structure
-
-
-
COFACTOR
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
Ca2+
-
may substitute for Mg2+, LPL-B
Co2+
-
may substitute for Mg2+
Co2+
-
may substitute for Mg2+, both LPL-A and LPL-B
Cu2+
-
may substitute for Mg2+, LPL-A
Fe2+
-
may substitute for Mg2+
Fe2+
-
may substitute for Mg2+, LPL-B
Fe3+
-
may substitute for Mg2+, LPL-B
Mg2+
-
Km-value 0.15 mM
Mg2+
-
required, both LPL-A and LPL-B
MgCl2
Q9HKT1, Q9HKT2
assay in the presence of; assay in the presence of
Mn2+
-
may substitute for Mg2+
Mn2+
-
may substitute for Mg2+, both LPL-A and LPL-B
Ni2+
-
may substitute for Mg2+
Ni2+
-
may substitute for Mg2+, both LPL-A and LPL-B
Zn2+
-
may substitute for Mg2+, both LPL-A and LPL-B
INHIBITORS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
6-seleno-octanoate
-
0.28 mM, complete inhibition, reactivation by reducing agents
8-bromooctanoate
-
inhibition of isoform LipL1 activity, in vitro growth arrest of Plasmodium falciparum
-
8-thiooctanoate
-
0.28 mM, 50% inhibition, reactivation by reducing agents
Cu2+
-
0.13 mM, 50% inhibition
additional information
-
not inhibitory: octanoyl-CoA
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
ATP
Q9HKT1, Q9HKT2
assay in the presence of; assay in the presence of
KM VALUE [mM]
KM VALUE [mM] Maximum
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
1.6
-
6-thio-octanoate
-
pH 7.0, 30C
0.0011
-
apoH-protein
-
mutant S72A, pH 7.0, 37C
-
0.0012
-
apoH-protein
-
wild-type, pH 7.0, 37C
-
0.0155
-
apoH-protein
-
mutant R140A, pH 7.0, 37C
-
0.0019
-
ATP
-
pH 7.0, 30C
0.0137
-
ATP
-
mutant R140A, pH 7.0, 37C
0.0158
-
ATP
-
wild-type, pH 7.0, 37C
0.295
-
ATP
-
mutant S72A, pH 7.0, 37C
0.0016
-
DL-lipoic acid
-
pH 7.0, 30C
0.0017
-
DL-lipoic acid
-
pH 7.0, 30C
0.2
-
LplA acceptor peptide 1
-
-
-
13.32
-
LplA acceptor peptide 2
-
-
-
0.004
-
Octanoic acid
-
mutant S221P
0.0104
-
Octanoic acid
-
mutant V19L
0.2143
-
Octanoic acid
-
wild type enzyme
0.00055
-
R-lipoic acid
-
mutant R140A, pH 7.0, 37C
0.0045
-
R-lipoic acid
-
wild-type, pH 7.0, 37C
0.0089
-
R-lipoic acid
-
mutant S72A, pH 7.0, 37C
0.013
-
selenolipoic acid
-
pH 7.0, 30C
TURNOVER NUMBER [1/s]
TURNOVER NUMBER MAXIMUM[1/s]
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
0.028
-
apoH-protein
-
mutant R140A, pH 7.0, 37C
-
0.212
-
apoH-protein
-
mutant S72A, pH 7.0, 37C
-
0.258
-
apoH-protein
-
wild-type, pH 7.0, 37C
-
0.0084
-
ATP
-
mutant R140A, pH 7.0, 37C
0.133
-
ATP
-
wild-type, pH 7.0, 37C
0.16
-
ATP
-
mutant S72A, pH 7.0, 37C
0.22
-
LplA acceptor peptide 2
-
-
-
0.0042
-
Octanoic acid
-
mutant S221P
0.0266
-
Octanoic acid
-
mutant V19L
0.04
-
Octanoic acid
-
wild type enzyme
0.006
-
R-lipoic acid
-
mutant R140A, pH 7.0, 37C
0.178
-
R-lipoic acid
-
mutant S72A, pH 7.0, 37C
0.219
-
R-lipoic acid
-
wild-type, pH 7.0, 37C
kcat/KM VALUE [1/mMs-1]
kcat/KM VALUE [1/mMs-1] Maximum
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
1.65e-05
-
LplA acceptor peptide 2
-
-
0
0.186
-
Octanoic acid
-
wild type enzyme
14763
1.05
-
Octanoic acid
-
mutant S221P
14763
2.55
-
Octanoic acid
-
mutant V19L
14763
SPECIFIC ACTIVITY [µmol/min/mg]
SPECIFIC ACTIVITY MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
11.5
-
-
37C
27.6
-
-
pH 7.0, 30C
additional information
-
-
Vmax of mutant S221P is 0.0025 nmol/min; Vmax of mutant V19L is 0.016 nmol/min; Vmax of wild type enzyme is 0.024 nmol/min
pH OPTIMUM
pH MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
7
-
-
assay at
7
-
-
assay at
pH RANGE
pH RANGE MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
5.5
8
-
more than 70% of maximum activity within this range
TEMPERATURE OPTIMUM
TEMPERATURE OPTIMUM MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
0
-
-
assay at
25
-
Q9HKT1, Q9HKT2
assay at; assay at
37
-
-
assay at
pI VALUE
pI VALUE MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
5.8
-
-
isoelectric focusing
6
-
-
calculated
SOURCE TISSUE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
SOURCE
-
developing, moderate expression, Northern blot
Manually annotated by BRENDA team
-
less abundant, Northern blot
Manually annotated by BRENDA team
-
developing seed, increasing level at 3 to 5 days after pollination followed by decreasing for 15 days, Northern blot
Manually annotated by BRENDA team
-
northern blot
Manually annotated by BRENDA team
additional information
-
in intraerythrocytic parasites, enzyme catalyzes incorporation of lipoate to mitochondrial proteins
Manually annotated by BRENDA team
additional information
-
transcript levels not influenced by reduced temperature (4C), high salt (100 mM NaCl) or 72 h treatment with 100 micromolar kinetin, 100 micromolar gibberellic acid or 50 micromolar alpha-naphthalene acetic acid
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
-
in intraerythrocytic parasites, enzyme catalyzes incorporation of lipoate to mitochondrial proteins
Manually annotated by BRENDA team
PDB
SCOP
CATH
ORGANISM
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Thermoplasma acidophilum (strain ATCC 25905 / DSM 1728 / JCM 9062 / NBRC 15155 / AMRC-C165)
Thermoplasma acidophilum (strain ATCC 25905 / DSM 1728 / JCM 9062 / NBRC 15155 / AMRC-C165)
Thermoplasma acidophilum (strain ATCC 25905 / DSM 1728 / JCM 9062 / NBRC 15155 / AMRC-C165)
Thermoplasma acidophilum (strain ATCC 25905 / DSM 1728 / JCM 9062 / NBRC 15155 / AMRC-C165)
Thermoplasma acidophilum (strain ATCC 25905 / DSM 1728 / JCM 9062 / NBRC 15155 / AMRC-C165)
Thermoplasma acidophilum (strain ATCC 25905 / DSM 1728 / JCM 9062 / NBRC 15155 / AMRC-C165)
MOLECULAR WEIGHT
MOLECULAR WEIGHT MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
11000
-
Q9HKT1, Q9HKT2
SDS-PAGE, molecular weight of HisCTD fusion protein
32000
-
Q9HKT1, Q9HKT2
SDS-PAGE, molecular weight of HisLplA fusion protein
41100
-
-
LplA-LplB complex, gel filtration
41700
-
-
gel filtration
SUBUNITS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
?
-
x * 47000, SDS-PAGE, both LPL-A and LPL-B
?
-
x * 38000, calculated
heterodimer
-
1 * 30800 + 1 * 11200, LplA-LplB complex, gel filtration
monomer
-
1 * 38000, SDS-PAGE
Crystallization/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
alone and in complex with lipoic acid
-
structural models of LplA1 and LplA2 based on Escherichia coli (PDB: 1X2H) and Streptococcus pneumoniae (PDB: 1VQZ) LplA proteins, conserved lipoyl-AMP binding pocket, LplA1: regions of clustered negatively charged residues, overall electrostatic potential of -13, LplA2: overall electrostatic potential of -5
-
structural comparison of lipoate-protein ligase with cysteine/lysine dyad acyltransferase LipB shows conserved structural and sequence active-site features, but 4-phosphopantheine-bound octanoic acid recognition is a specific property of cysteine/lysine dyad acyltransferase
-
as apo form, in complex with ATP, and as lipoyl-AMP-complex. Lipoyl-AMP is bound deeply in the bifurcated pocket and adopts a U-shaped conformation
-
OXIDATION STABILITY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
slow oxidation to an inactive form containing at least one intramolecular disulfide bond, reactivation by reducing agent
-
663919
STORAGE STABILITY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
-20C, stable for several months
-
4C, slow oxidation to an inactive form containing at least one intramolecular disulfide bond, reactivation by reducing agent
-
Purification/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
; Ni-NTA column chromatography, ammonium sulfate precipitation gel filtration
-
both LPL-A and LPL-B, partial
-
Ni2+-NTA-agarose resin column chromatography and POROS HQ 20 column chromatography
-
using His-tag affinity purification; using His-tag affinity purification
Q9HKT1, Q9HKT2
Cloned/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
expressed in Escherichia coli
-
in Escherichia coli
-
LplA1: in pAM401 for complementation in Listeria monocytogenes mutant strains DP-L4263 and 10403S, in ptac85 for IPTG-inducible expression in Escherichia coli wild-type strain JK1 or strain TM131 lacking endogenous LplA, LplA2: from bacterial genome (10403S) in pKSV7 for allelic exchange (homologous recombination), in ptac85 for IPTG-inducible expression in Escherichia coli wild-type strain JK1 or strain TM131 lacking endogenous LplA
-
from cDNA library in pTrcHis2A for expression and functional complementation in Escherichia coli strain TM137 (lplA/lipB null mutant of strain JK1)
-
expressed in Escherichia coli
-
expressed in Escherichia coli as a His-tagged fusion protein; expressed in Escherichia coli as a His-tagged fusion protein
Q9HKT1, Q9HKT2
ENGINEERING
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
D122A
-
D122A mutation results in a marked reduction in the overall, lipoate adenylation, and lipoate transfer reaction activities (0.14, 4, and 4% of those of wild type, respectively)
E116A/E312K/L328F
-
mutations allow a LipB knockout strain to grow on a glucose minimal medium
F15S/T101A/S114I
-
mutations allow a LipB knockout strain to grow on a glucose minimal medium
F35L/V113I
-
mutations allow a LipB knockout strain to grow on a glucose minimal medium
G76S
-
substitution in LplA ligase gene, is identical to slr1 selenolipoate restistance mutation
H149A
-
mutations does not cause a significant reduction in three reaction activities (overall, lipoate adenylation, and lipoate transfer reaction activities), Km value for ATP and lipoic acid increases to 15 and 5.8fold, respectively, relative to those of wild-type
N121A
-
N121A affects only the lipoate adenylation activity and consequently the overall reaction activity (1.4 and 0.19% of those of wild-type, respectively) but retains a significant lipoate transfer activity (24.2%)
R140A
-
12fold increase in Km-value for apoH-protein
R58L/H79N
-
mutations allow a LipB knockout strain to grow on a glucose minimal medium
S221P
-
mutant with reduced affinity to octanoic acid; strain FH27, referred as lplA11, mutation allows a LipB knockout strain to grow on a glucose minimal medium
S72A
-
great increase in Km-value for ATP
S72A
-
mutations does not cause a significant reduction in three reaction activities (overall, lipoate adenylation, and lipoate transfer reaction activities), Km value for ATP and lipoic acid increases to 28 and 2.3fold, respectively, relative to those of wild-type
S8T/N63K/F78Y/A110T
-
mutations allow a LipB knockout strain to grow on a glucose minimal medium
K133A
-
K133A mutation almost completely abolishes the overall reaction activity (0.01% of that of wild type) and causes marked reduction in lipoate adenylation and lipoate transfer activities (0.2 and 2.5% of that of wild type, respectively)
additional information
-
lipB mutant strain, grows well when supplemented with octanoate in place of lipoate
additional information
-
lplA null mutants display no growth defect unless combined with lipA or lipB lipoate synthesis mutations
additional information
-
enzyme null mutant, normal transport of lipoic acid, but severe defect in incorporation and utilization of exogenously supplied lipoic acid and lipoic acid analogues. Strain is highly resistant to selenolipoate
V19L
-
mutant with reduced affinity to octanoic acid; strain FH26, referred as lplA10, mutation allows a LipB knockout strain to grow on a glucose minimal medium
additional information
-
knockout of Plasmodium falciparum LplA1 failed because the protein is essential for growth of parasite
additional information
Q9HKT1, Q9HKT2
LplA and CTD encoding genes are expressed as fusion proteins in Escherichia coli by omitting the stop codon of lplA and the start codon of ctd. Fusion protein is shown to be catalytically active; LplA and CTD encoding genes are expressed as fusion proteins in Escherichia coli by omitting the stop codon of lplA and the start codon of ctd. Fusion protein is shown to be catalytically active
Renatured/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
reactivation of inactive form containing at least one intramolecular disulfide bond, or of enzyme inactivated by substrate-analogues, by reducing agent
-
APPLICATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
drug development
-
LplA1, but not LplA2, required for intracellular replication and/or virulence of Listeria monocytogenes
molecular biology
-
dependency of pathogen Listeria monocytogenes on LplA1 dictated by availability of host lipoyl substrates as alternative lipoate source