Information on EC 2.3.1.129 - acyl-[acyl-carrier-protein]-UDP-N-acetylglucosamine O-acyltransferase

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

EC NUMBER
COMMENTARY hide
2.3.1.129
-
RECOMMENDED NAME
GeneOntology No.
acyl-[acyl-carrier-protein]-UDP-N-acetylglucosamine O-acyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
(R)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-N-acetyl-alpha-D-glucosamine = an [acyl-carrier protein] + UDP-3-O-[(3R)-3-hydroxytetradecanoyl]-N-acetyl-alpha-D-glucosamine
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Acyl group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
lipid A biosynthesis
-
-
lipid IVA biosynthesis
-
-
Lipopolysaccharide biosynthesis
-
-
Metabolic pathways
-
-
SYSTEMATIC NAME
IUBMB Comments
(R)-3-hydroxytetradecanoyl-[acyl-carrier protein]:UDP-N-acetylglucosamine 3-O-(3-hydroxytetradecanoyl)transferase
Involved with EC 2.4.1.182 (lipid-A-disaccharide synthase) and EC 2.7.1.130 (tetraacyldisaccharide 4'-kinase) in the biosynthesis of the phosphorylated glycolipid, Lipid A, in the outer membrane of Escherichia coli.
CAS REGISTRY NUMBER
COMMENTARY hide
105843-69-4
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SM105
-
-
Manually annotated by BRENDA team
Escherichia coli strains JB1104
strains JB1104
-
-
Manually annotated by BRENDA team
W3106
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
strain O35E, gene lpxA
SwissProt
Manually annotated by BRENDA team
strain O35E, gene lpxA
SwissProt
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
gene lpxA
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
metabolism
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
(3R)-3-hydroxylauroyl-methylphosphopantetheine + UDP-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
methylphosphopantetheine + UDP-3-N-((3R)-3-hydroxylauroyl)-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
show the reaction diagram
synthetic (3R)-3-hydroxylauroyl-methylphosphopantetheine is an excellent alternative donor substrate for LiLpxA
-
-
?
(R)-3-hydroxydecanoyl-[acyl-carrier protein] + UDP-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
[acyl-carrier protein] + UDP-3-N-(3-hydroxydecanoyl)-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
show the reaction diagram
-
-
-
-
?
(R)-3-hydroxydodecanoyl-[acyl-carrier protein] + UDP-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
[acyl-carrier protein] + UDP-3-N-(3-hydroxydodecanoyl)-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
show the reaction diagram
(R)-3-hydroxydodecanoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxydodecanoyl)-N-acetylglucosamine
show the reaction diagram
-
-
-
-
?
(R)-3-hydroxyhexadecanoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxyhexadecanoyl)-N-acetylglucosamine
show the reaction diagram
-
-
-
-
?
(R)-3-hydroxylauroyl-[acyl-carrier protein] + UDP-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
[acyl-carrier protein] + UDP-3-N-((3R)-3-hydroxylauroyl)-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
show the reaction diagram
LiLpxA is highly selective for (R)-3-hydroxylauroyl-[acyl-carrier protein] and UDP-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
-
-
?
(R)-3-hydroxymyristoyl-[acyl carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-((3R)-3-hydroxymyristoyl)-alpha-D-glucosamine
show the reaction diagram
first step in lipid A biosynthesis
-
-
?
(R)-3-hydroxymyristoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxymyristoyl)-N-acetylglucosamine
show the reaction diagram
first step in lipid A biosynthesis
-
-
?
(R)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
[acyl-carrier protein] + UDP-3-N-(3-hydroxytetradecanoyl)-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
show the reaction diagram
(R)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxytetradecanoyl)-N-acetylglucosamine
show the reaction diagram
(R)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-N-butyrylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxytetradecanoyl)-N-butyrylglucosamine
show the reaction diagram
-
transacylation at 8% the rate of the reaction with UDP-N-acetylglucosamine
-
-
?
(R)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-N-propionylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxytetradecanoyl)-N-propionylglucosamine
show the reaction diagram
-
transacylation at 22% the rate of the reaction with UDP-N-acetylglucosamine
-
-
?
(R,S)-3-hydroxydodecanoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxydodecanoyl)-N-acetylglucosamine
show the reaction diagram
(R,S)-3-hydroxyhexadecanoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxyhexadecanoyl)-N-acetylglucosamine
show the reaction diagram
(R,S)-3-hydroxylauroyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxylauroyl)-N-acetylglucosamine
show the reaction diagram
(R,S)-3-hydroxymyristoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxymyristoyl)-N-acetylglucosamine
show the reaction diagram
(R,S)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxytetradecanoyl)-N-acetylglucosamine
show the reaction diagram
(S)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxytetradecanoyl)-N-acetylglucosamine
show the reaction diagram
-
transacylation at 7% the rate of (R)-enantiomer
-
-
?
an (R)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-N-acetyl-alpha-D-glucosamine
an [acyl-carrier protein] + UDP-3-O-(3-hydroxytetradecanoyl)-N-acetyl-alpha-D-glucosamine
show the reaction diagram
decanoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-decanoyl-N-acetylglucosamine
show the reaction diagram
-
-
-
-
?
myristoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-myristoyl-N-acetylglucosamine
show the reaction diagram
-
higher specific activity than to (R,S)-3-hydroxymyristoyl-[acyl-carrier-protein]
-
-
?
R-3-hydroxydecanoyl-[acyl carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-o-hydroxydecanoyl-N-acetylglucosamine
show the reaction diagram
first step in lipid A biosynthesis
-
-
?
additional information
?
-
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
(R)-3-hydroxydodecanoyl-[acyl-carrier protein] + UDP-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
[acyl-carrier protein] + UDP-3-N-(3-hydroxydodecanoyl)-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
show the reaction diagram
-
first step in lipid A biosynthesis pathway, lipid A is an unusual variant in this species containing 2,3-diamino-2,3-dideoxy-D-glucopyranose units
-
-
?
(R)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
[acyl-carrier protein] + UDP-3-N-(3-hydroxytetradecanoyl)-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
show the reaction diagram
(R)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-N-acetylglucosamine
[acyl-carrier protein] + UDP-3-O-(3-hydroxytetradecanoyl)-N-acetylglucosamine
show the reaction diagram
an (R)-3-hydroxytetradecanoyl-[acyl-carrier protein] + UDP-N-acetyl-alpha-D-glucosamine
an [acyl-carrier protein] + UDP-3-O-(3-hydroxytetradecanoyl)-N-acetyl-alpha-D-glucosamine
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
-
not as effective as Mg2+
KCl
-
slight activation at 0.25 M
LiCl
-
slight activation at 0.25 M
Mg2+
-
is required for stabilization of substrate acyl-carrier protein in a helical conformation
Mn2+
-
not as effective as Mg2+
NaCl
-
slight activation at 0.25 M
sulfate
-
2 ions per asymmetrical unit
Tartrate
-
1 ions per asymmetrical unit
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
diethyldicarbonate
-
-
DL-3-hydroxymyristic acid
-
competitive versus TAMRA peptide
FITC-P920
-
i.e. FITC-(beta)SSGWMLDPIAGKWSRNH2, fluoresecent-labeled peptide
HM-UNAG peptide
-
-
-
HMA peptide
-
-
-
KCl
-
0.5 M
LiCl
-
0.5 M
myristoyl-[acyl-carrier protein]
NaCl
-
0.5 M
P920
-
i.e. SSGWMLDPIAGKWSR
-
peptide 920
Phenylglyoxal
-
-
pyridoxal 5'-phosphate/sodium borohydride
-
-
RJPXD31
-
i.e. QHFMVPDINDMQ-NH2
RJPXD33
-
i.e. TNLYMLPKWDIP-NH2, a peptide identified from a phage-bound random peptide library screen uing Escherichia oli strain XL-1 Blue, binds to UDP-3-O-(R-3-hydroxyacyl)GlcN N-acyltransferase, LpxD, and UDP-N-acetylglucosamine acyltransferase, LpxA. RJPXD33 binds to LpxA in a competitive fashion with P920
TAMRA peptide
-
ability of the TAMRA-peptide to inhibit the catalytic activity
-
Triton X-100
-
-
UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine
-
competitive versus TAMRA peptide
UDP-N-acetylglucosamine
-
competitive versus TAMRA peptide
UNAG peptide
-
-
-
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0016 - 0.005
(R)-3-hydroxytetradecanoyl-[acyl-carrier protein]
0.004
3R-3-hydroxylauroyl-methylphosphopantetheine
-
0.099 - 15
UDP-N-acetylglucosamine
additional information
additional information
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
7.17 - 10.5
UDP-N-acetylglucosamine
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00005
peptide 920
additional information
additional information
inhibition kinetics
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.05
DL-3-hydroxymyristic acid
Escherichia coli
-
pH 7.0, 22C
0.00014 - 0.000238
HM-UNAG peptide
0.047 - 0.063
HMA peptide
0.00006 - 0.000923
peptide 920
0.019
RJPXD33
Escherichia coli
-
pH 7.5, 30C
0.00015
TAMRA peptide
Escherichia coli
-
forward reaction, pH 7.0, 22C
-
0.0002
UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine
Escherichia coli
-
pH 7.0, 22C
6
UDP-N-acetylglucosamine
Escherichia coli
-
pH 7.0, 22C
5.8 - 6.6
UNAG peptide
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.00000002
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO15
0.00000014
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate, pTO14
0.00000028
-
S)-3-hydroxydodecanoyl-[acyl-carrier-protein] as substrate, pTO15
0.00000034
-
S)-3-hydroxydodecanoyl-[acyl-carrier-protein] as substrate, pTO14
0.00000072
-
S)-3-hydroxyhexadecanoyl-[acyl-carrier-protein]
0.000001
-
S)-3-hydroxydodecanoyl-[acyl-carrier-protein] as substrate, pTO202; S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO9
0.0000011
-
S)-3-hydroxymyristoyl-[acyl-carrier-protein] as substrate
0.0000015
-
decanoyl-[acyl-carrier-protein] as substrate
0.000003
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate, pTO201
0.0000032
-
S)-3-hydroxylauroyl-[acyl-carrier-protein] as substrate
0.000006
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate, pTO10 and pTO105
0.000007
-
S)-3-hydroxydodecanoyl-[acyl-carrier-protein] as substrate, pTO10
0.00001
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO202
0.000011
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate, pTO15
0.000014
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO14
0.000015
-
S)-3-hydroxydodecanoyl-[acyl-carrier-protein]
0.000019
-
S)-3-myristoyl-[acyl-carrier-protein] as substrate
0.000022
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate, pTO105
0.000023
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate, pTO103
0.000024
-
S)-3-hydroxydodecanoyl-[acyl-carrier-protein] as substrate, pTO201
0.00003
-
SM 101
0.000038
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate, pTO202
0.00004
-
S)-3-hydroxydodecanoyl-[acyl-carrier-protein] as substrate, pTO9
0.000054
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate, pTO1
0.000055
-
S)-3-hydroxymyristoyl-[acyl-carrier-protein] as substrate
0.00006
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO101
0.000067
-
S)-3-hydroxymyristoyl-[acyl-carrier-protein] as substrate
0.000068
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO104
0.00007
0.000079
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate, pTO101
0.000087
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO103
0.0002
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO102
0.000247
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate
0.000347
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO10
0.00035
0.00047
-
S)-3-hydroxydodecanoyl-[acyl-carrier-protein]
0.00053
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO102
0.00059
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein]
0.00079
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein]
0.00087
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein]
0.0011
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein]
0.0012
0.00139
-
S)-3-hydroxylauroyl-[acyl-carrier-protein] as substrate
0.0015
-
SM 105
0.00325
-
S)-3-hydroxylauroyl-[acyl-carrier-protein] as substrate
0.011
-
S)-3-hydroxydecanoyl-[acyl-carrier-protein] as substrate
0.074
-
S)-3-hydroxytetradecanoyl-[acyl-carrier-protein] as substrate, pTO1
3
using synthetic (3R)-3-hydroxylauroyl-methylphosphopantetheine + UDP-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose as a substrate the specific activity is comparable to substrates (3R)-3-hydroxylauroyl-[acyl-carrier protein] + UDP-2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.8 - 8.2
-
broad
7
-
assay at, binding measurements are performed at pH 8.0
7.5
-
activity assay at
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
22
-
assay at room temperature
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Acinetobacter baumannii (strain TCDC-AB0715)
Acinetobacter baumannii (strain TCDC-AB0715)
Bacteroides fragilis (strain ATCC 25285 / DSM 2151 / JCM 11019 / NCTC 9343)
Bacteroides fragilis (strain ATCC 25285 / DSM 2151 / JCM 11019 / NCTC 9343)
Burkholderia thailandensis (strain E264 / ATCC 700388 / DSM 13276 / CIP 106301)
Campylobacter jejuni subsp. jejuni serotype O:2 (strain ATCC 700819 / NCTC 11168)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Helicobacter pylori (strain ATCC 700392 / 26695)
Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai (strain 56601)
Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai (strain 56601)
Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai (strain 56601)
Pseudomonas aeruginosa (strain PA7)
Pseudomonas aeruginosa (strain PA7)
Pseudomonas aeruginosa (strain PA7)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
28000
-
SDS-PAGE
90000
-
gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
-
x * 27000, calculated from nucleotide sequence
dimer or trimer
-
alpha2, 2 * 30000, SDS-PAGE
homotrimer
trimer
additional information
-
enzyme structure contains a left-handed parallel beta-helix motif
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified recombinant LpxA, sitting drop vapor diffusion method, mixing of AtLpxA at 18 mg/mL in 10 mM potassium phosphate buffer, pH 7.0, 200 mM KCl, and 20% glycerol, in a 1:1 ratio with precipitant solution containing 0.5 M ammonium sulfate, 0.1 M sodium citrate, pH 5.6, and 1.0 M lithium sulfate monohydrate, equilibration at 20C, 1-2 days, X-ray diffraction structure determination and analysis at 2.1 A resolution
by the sitting-drop, vapour-diffusion method using 0.00l ml of protein solution; LpxA in complex with UDP-GlcNAc, modelling
by using the hanging drop vapor diffusion method, in the presence of a 25-fold molar excess of either UDP-3-O-(R-3-hydroxymyristoyl)-GlcNAc or UDP-3-O-(R-3-hydroxydecanoyl)-GlcNAc, structures show how LpxA selects for 14-carbon R-3-hydroxyacyl chains and reveal two modes of UDP binding
-
purified enzyme LpxA in a complex with inhibitor peptide 920, 20 mg/ml protein in solution with a 25fold molar excess of peptide 920 of 12.5 mM, crystal growth at 18C, hanging drop vapor diffusion method, mixing of 0.002 ml protein solution with 0.002 ml of 0.81.8 M phosphate buffer, pH 6.36.9, and 30-35% DMSO, about 2 weeks, X-ray diffraction structure determination and analysis at 1.8 A resolution, molecular replacement using crystal structure PDB ID code 1LXA, determined at 2.6 A resolution
crystallized at 297 K using (NH4)2SO4 and Na/K tartrate as precipitants in the presence of a detergent, space group: P6322 with unit cell-parameters: a = b = 90.69, and c = 148.20 ANG
-
purified recombinant, C-terminally eight-residue-tagged enzyme, X-ray diffraction structure analysis at 2.1 A resolution, modeling of the enzyme complexed with [acyl-carrier protein]
-
crystal structures of free LiLpxA and its complexes with its product UDP-3-N-((R)-3-hydroxylauroyl)-GlcNAc3N and with its substrate (R)-3-hydroxylauroyl-methylphosphopantetheine are presented. The selectivity of LiLpxA for UDPGlcNAc3N may be explained by the orientation of the backbone carbonyl group of Q68, which differs by 82 from the corresponding Q73 carbonyl group in Escherichia coli LpxA. This arrangement provides an extra hydrogen-bond acceptor for the 3-NH2 group of UDP-GlcNAc3N in LiLpxA. The R-3-hydroxylauroyl selectivity of LiLpxA is explained by the position of the K171 side chain, which limits the length of the acyl-chain-binding groove. H120 functions as a catalytic base
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60
-
20 min stable
100
-
inactivation after 10 min
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
bovine serum albumin prevents denaturation
-
freeze-thawing inactivates
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-80C, flash-frozen in liquid nitrogen, at least 3 months
-
4C, several days
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ion-exchange-chromatography and gel filtration
partial, overproducing strain MC1061/pSR1
-
recombinant LpxA from Echerichia coli strain BL21-AI/pET24a by Reactive Green 19 affinity chromatography, dialysis, and ion exchange chromatography
-
recombinant LpxA, excluding the region encoding the predicted signal peptide (residues 2-32), from Escherichia coli strain C41(DE3) by Reactive Green 19 affinity chromatography and gel filtration to homogeneity
using Ni-NTA chromatography and gel filtration. Around 15 mg of purified protein is obtained from 1 l of bacterial culture
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherchia coli as a His-tagged fusion protein
-
expressed in Escherichia coli
expression in Corynebacterium glutamicum
-
expression in Escherichia coli
gene lpxA, cloning from strain strain MG1655, subcloning in strain XL-1 Blue, expression in Escherichia coli strain BL21-AI/pET24a
-
gene lpxA, DNA and amino acid sequence determination and analysis, expression in Escherichia coli strain BL21(DE3)
gene lpxA, DNA and amino acid sequence determination, subcloning in Escherichia coli, wild-type and mutant strains expression analysis
gene lpxA, excluding the region encoding the predicted signal peptide (residues 2-32), expression of the plant enzyme in Escherichia coli strain C41(DE3). Functional expression of AtlpxA in and complementation of the constructed lpxA Escherichia coli knockout mutant strains
gene lpxA, expression in Escherichia coli strain BL21(DE3)
-
gene lpxA, expression in strain BL21(DE3)
-
gene lpxA, five AtLpxC orthologues located in a 68-kb region on chromosome one, DNA and amino acid sequence determination and analysis, sequence comparison
overexpression in Escherichia coli
recombinant expresssion
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
G189S
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Escherichia coli strain SM101 is deficient in LpxA activity due to a G189S inactivating mutation. Enzymatic activity is restored when the mutant strain is transformed with a wild-type bearing plasmid is inactive
G52L/R58L
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residue tolerance in the beta-helical domain: mutation is tolerated
G52V/R58V
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residue tolerance in the beta-helical domain: mutation is tolerated
G52W/R58W
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residue tolerance in the beta-helical domain: mutation is tolerated
G57P/N51P
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residue tolerance in the beta-helical domain: mutation is not tolerated
H122A
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lower specific activity than wild-type
H122N
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lower specific activity than wild-type
H125N
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lower specific activity than wild-type
H144A
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lower specific activity than wild-type
H144N
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lower specific activity than wild-type
H160A
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lower specific activity than wild-type
H160F
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lower specific activity than wild-type
H53I/D59I
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residue tolerance in the beta-helical domain: mutation is tolerated
H53V/D59V
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residue tolerance in the beta-helical domain: mutation is tolerated
I20R
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mutation in the hydrophobic residue in the beta-helical core located in rung 2: LpxA is active
I2R
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mutation in the hydrophobic residue in the beta-helical core located in rung 1: LpxA is active
I38R
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mutation in the hydrophobic residue in the beta-helical core located in rung 3: LpxA is partially active
I56A
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mutation in the hydrophobic core of the beta-helical domain: LpxA activity is not significantly affected compared to wild-type
I56D/I62D
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residue tolerance in the beta-helical domain: mutation is not tolerated
I56E/I62E
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residue tolerance in the beta-helical domain: mutation is not tolerated
I56G
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mutation in the hydrophobic core of the beta-helical domain: LpxA activity is decreased compared to wild-type
I56G/I62G
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residue tolerance in the beta-helical domain: mutation is tolerated
I56H/I62H
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residue tolerance in the beta-helical domain: mutation is not tolerated
I56K/I62K
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residue tolerance in the beta-helical domain: mutation is not tolerated
I56N
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mutation in the hydrophobic core of the beta-helical domain: LpxA activity is decreased compared to wild-type
I56N/I62N
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residue tolerance in the beta-helical domain: mutation is tolerated
I56P/I62P
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residue tolerance in the beta-helical domain: mutation is not tolerated
I56Q
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mutation in the hydrophobic core of the beta-helical domain: LpxA activity is decreased compared to wild-type
I56Q/I62Q
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residue tolerance in the beta-helical domain: mutation is tolerated
I56R
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mutation in the hydrophobic core of the beta-helical domain: LpxA activity is completely abolished compared to wild-type; mutation in the hydrophobic residue in the beta-helical core located in rung 4: LpxA is inactive
I56R/I62R
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residue tolerance in the beta-helical domain: mutation is not tolerated
I56S/I62S
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residue tolerance in the beta-helical domain: mutation is tolerated
I56W/I62W
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residue tolerance in the beta-helical domain: mutation is not tolerated
I56Y/I62Y
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residue tolerance in the beta-helical domain: mutation is not tolerated
I86R
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mutation in the hydrophobic residue in the beta-helical core located in rung 5: LpxA is inactive; residue is located in the fifth rung of the beta-helical domain in the hydrophobic core of a beta helix, mutant is improperly folded, destabilized and its enzymatic activity is decreased. Mutant shows significant growth reduction after introduction into Escherichia coli strain SM101 bearing a defective LpxA gene (G189S), and under novobiocin supplementation
K55D/E61D
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residue tolerance in the beta-helical domain: mutation is tolerated
K55P/E61P
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residue tolerance in the beta-helical domain: mutation is not tolerated
K76A
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lower specific activity than wild-type
K76R
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lower specific activity than wild-type
P10A/P28A/P34A/P183A
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proline mutation at the turn region of the beta-helical domain: mutant shows lower activity compared to wild-type
P28A/P34A
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proline mutation at the turn region of the beta-helical domain: mutant shows greater activity than the P10A/P28A/P34A/P183A mutant
T54D/N60D
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residue tolerance in the beta-helical domain: mutation is not tolerated
T54E/N60E
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residue tolerance in the beta-helical domain: mutation is not tolerated
T54G/N60G
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residue tolerance in the beta-helical domain: mutation is tolerated
T54H/N60H
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residue tolerance in the beta-helical domain: mutation is not tolerated
T54K/N60K
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residue tolerance in the beta-helical domain: mutation is not tolerated
T54M/N60M
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residue tolerance in the beta-helical domain: mutation is tolerated
T54P/N60P
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residue tolerance in the beta-helical domain: mutation is not tolerated
T54Q/N60Q
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residue tolerance in the beta-helical domain: mutation is tolerated
T54R/N60R
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residue tolerance in the beta-helical domain: mutation is not tolerated
T54W/N60W
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residue tolerance in the beta-helical domain: mutation is not tolerated
T54Y/N60Y
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residue tolerance in the beta-helical domain: mutation is not tolerated
V111R
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mutation in the hydrophobic residue in the beta-helical core located in rung 6: LpxA is inactive
V129R
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mutation in the hydrophobic residue in the beta-helical core located in rung 7: LpxA is inactive
Y66F/Y77F/Y219F/Y223F/Y243H
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all but one tyrosine residues are mutated. Mutant shows growth similar to wild-type after introduction into Escherichia coli strain SM101 bearing a defective LpxA gene (G189S), and under novobiocin supplementation
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
drug development
medicine
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structures helps to design new inhibitors that target LpxA, antibiotics
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