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Information on EC 6.3.2.43 - [amino group carrier protein]-L-2-aminoadipate ligase

for references in articles please use BRENDA:EC6.3.2.43
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EC Tree
IUBMB Comments
The enzymes from the thermophilic bacterium Thermus thermophilus and the thermophilic archaea Sulfolobus acidocaldarius and Sulfolobus tokodaii protect the amino group of L-2-aminoadipate by conjugation to the carrier protein LysW. This reaction is part of the lysine biosynthesis pathway in these organisms.
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UNIPROT: Q5JFW0
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The expected taxonomic range for this enzyme is: Bacteria, Archaea
Synonyms
alpha-aminoadipate-lysW ligase lysX, LysX, More, RimK-related lysine biosynthesis protein, Saci_0754, Tk0278, TkLysX/ArgX, TtLysX, [lysine-biosynthesis-protein LysW]-C-terminal-L-glutamate:L-2-aminoadipate ligase (ADP-forming), [lysine-biosynthesis-protein LysW]-L-2-aminoadipate ligase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
RimK-related lysine biosynthesis protein
UniProt
[lysine-biosynthesis-protein LysW]-C-terminal-L-glutamate:L-2-aminoadipate ligase (ADP-forming)
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[lysine-biosynthesis-protein LysW]-L-2-aminoadipate ligase
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additional information
cf. EC 6.3.2.60
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + an [amino group carrier protein]-C-terminal-L-glutamate + L-2-aminoadipate = ADP + phosphate + an [amino group carrier protein]-C-terminal-N-(1,4-dicarboxybutyl)-L-glutamine
show the reaction diagram
substrate-recognition mechanism of bifunctional TkLysX/ArgX
PATHWAY SOURCE
PATHWAYS
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SYSTEMATIC NAME
IUBMB Comments
[amino group carrier protein]-C-terminal-L-glutamate:L-2-aminoadipate ligase (ADP-forming)
The enzymes from the thermophilic bacterium Thermus thermophilus and the thermophilic archaea Sulfolobus acidocaldarius and Sulfolobus tokodaii protect the amino group of L-2-aminoadipate by conjugation to the carrier protein LysW. This reaction is part of the lysine biosynthesis pathway in these organisms.
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + an [amino group carrier protein]-C-terminal-L-glutamate + L-2-aminoadipate
ADP + phosphate + an [amino group carrier protein]-C-terminal-N-(1,5-dicarboxypentan-1-yl)-L-glutamine
show the reaction diagram
ATP + [lysine-biosynthesis-protein LysW]-C-terminal-L-glutamate + L-2-aminoadipate
ADP + phosphate + [lysine-biosynthesis-protein LysW]-C-terminal-gamma-(L-2-aminoadip-2-yl)-L-glutamate
show the reaction diagram
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?
ATP + [lysine-biosynthesis-protein LysW]-C-terminal-L-glutamate + L-glutamate
ADP + phosphate + [lysine-biosynthesis-protein LysW]-C-terminal-gamma-(L-glutam-2-yl)-L-glutamate
show the reaction diagram
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ATP + [LysW mutant E42R]-C-terminal-L-glutamate + L-2-aminoadipate
ADP + phosphate + [LysW mutant E42R]-C-terminal-N-(1,5-dicarboxypentan-1-yl)-L-glutamine
show the reaction diagram
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ATP + [LysW]-C-terminal-L-glutamate + L-2-aminoadipate
ADP + phosphate + [LysW]-C-terminal-N-(1,4-dicarboxybutan-1-yl)-L-glutamine
show the reaction diagram
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additional information
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purified recombinant TK0278 recognizes alpha-aminoadipate (AAA) and glutamate as substrates and ligated both compounds with TK0279, an amino group carrier protein LysW in this microorganism, in an ATP-dependent manner
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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
ATP + an [amino group carrier protein]-C-terminal-L-glutamate + L-2-aminoadipate
ADP + phosphate + an [amino group carrier protein]-C-terminal-N-(1,5-dicarboxypentan-1-yl)-L-glutamine
show the reaction diagram
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?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Mg2+
required, nine Mg2+ ions per dimeric enzyme tetramer
additional information
LysW contains a Zn2+
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.016
substrate L-glutamate, pH not specified in the publication, temperature not specified in the publication
0.021
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
Pyrococcus kodakaraensis strain KOD1
UniProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
proposal of a mechanism for substrate recognition and its relationship with molecular evolution among LysX family proteins, which have different substrate specificities
metabolism
the enzyme is part of the lysine biosynthetic pathway in Thermococcus kodakarensis, overview. The gene cluster for lysine biosynthetic genes through AAA produces lysine with a LysW-mediated system. In this cluster, TK0279, TK0278, TK0276, TK0277, TK0275, and TK0274 are expected to be responsible for the conversion process from AAA to lysine through the LysW system as LysW-gamma-AAA synthetase, LysW-gamma-AAA kinase, LysW-gamma-AAA semialdehyde dehydrogenase, LysW-gamma-lysine aminotransferase, and LysW-gamma-lysine carboxypeptidase, respectively. LysW-gamma-AAA thus synthesized is transferred to subsequent biosynthetic enzymes to be converted to LysW-gamma-lysine by phosphorylation, reduction, and amination steps. In the final step, LysW-gamma-lysine is recognized by LysK, a carboxypeptidase, resulting in the release of lysine. The lysine biosynthetic enzymes of Thermococcus kodakarensis convert AAA/Glu to lysine/ornithine
physiological function
additional information
the modification by TK0278 and successive phosphorylation by TK0276 occurr at the C-terminus of TkLysW. Residue Tyr175 in TkLysX/ArgX plays a critical role in the recognition of glutamate. Residues Thr196, Asn250, and Ala251 are involved in the recognition of the delta-carboxyl group of AAA. The Tyr residue at strand beta10, the Tyr residue at strand beta11, and the Thr residue at the large loop as the additional key residues that define substrate specificity in LysX family proteins
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION?
Q5JFW0_THEKO
Thermococcus kodakarensis (strain ATCC BAA-918 / JCM 12380 / KOD1)
273
0
30488
TrEMBL
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SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
tetramer
the crystal asymmetric unit contains two TkLysX/ArgX tetramers, two intact TkLysW monomers each with one zinc atom, four partial TkLysW, eight ADP molecules, five phosphate ions, two sulfate ions, nine magnesium atoms, one free AAA molecule, and 628 water molecules, binding structure, overview
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
in presence of carrier protein LysW and AMP-PNP, MgSO4, and L-2-aminoadipate or glutamate, to 2.18 A resolution
purified recombinant nontagged enzyme TkLysX/ArgX in complex with [LysW]-C-terminal-N-(1,5-dicarboxypentan-1-yl)-L-glutamine ((TkLysX/ArgXx02LysW-gamma-AAA)), which corresponds to the structure of the post-reaction state of TkLysX/Arg, hanging drop vapor diffusion method, mixing of 7.5 mg/ml protein and 2.5 mg/ml TkLysW in 10 mM AMP-PNP, 10 mM AAA, or 50 mM glutamate and 10 mM MgSO4 with precipitation solution containing containing 20% v/v 2-methyl-2,4-pentanediol, 6.6% w/v PEG-3350, 0.066 M imidazole, pH 6.5, and 0.132 M ammonium sulfate, 20°C, X-ray diffraction structure determination and analysis at 2.18 A resolution. Molecular replacement. The asymmetric unit contains two TkLysX/ArgX tetramers, two intact TkLysW monomers each with one zinc atom, four partial TkLysW, eight ADP molecules, five phosphate ions, two sulfate ions, nine magnesium atoms, one free AAA molecule, and 628 water molecules. Good crystals containing TkLysW-gamma-Glu are not obtained
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
I185Y
site-directed mutagenesis, the mutant exhibits an apparent substrate preference for glutamate
N250G
site-directed mutagenesis, the mutant exhibits an apparent substrate preference for glutamate
S251F
site-directed mutagenesis, the mutant exhibits an apparent substrate preference for glutamate
Y175I
site-directed mutagenesis, the mutant exhibits an apparent substrate preference for glutamate
Y175I/I185Y/N250G/S251F
mutant exhibits an apparent substrate preference for glutamate
additional information
the N250G/A251F mutant is not produced as a soluble protein
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant C-terminally His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21-CodonPlus (DE3)-RIL by heat treatment at 80°C for 20 min and nickel affinity chromatography. Recombinant nontagged enzyme from Escherichia coli strain BL21(DE3) by heat treatment at 80°C for 20 min, ammonium sulfate fractionation, hydrophobic interaction chromatography, ultrafiltration, and gel filtration
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
gene TK0278, recombinant expression of C-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21-CodonPlus (DE3)-RIL, recombinant expression of nontagged enzyme in Escherichia coli strain BL21(DE3)
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Yoshida, A.; Tomita, T.; Atomi, H.; Kuzuyama, T.; Nishiyama, M.
Lysine biosynthesis of Thermococcus kodakarensis with the capacity to function as an ornithine biosynthetic system
J. Biol. Chem.
291
21630-21643
2016
Thermococcus kodakarensis (Q5JFW0), Thermococcus kodakarensis ATCC BAA-918 (Q5JFW0), Thermococcus kodakarensis JCM 12380 (Q5JFW0)
Manually annotated by BRENDA team