Literature summary for 2.3.2.3 extracted from

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 (2016), J. Biol. Chem., 291, 21630-21643 .

Activating Compound

Activating Compound	Comment	Organism	Structure
ADP	1 ADP is located in the active site of the enzyme	Thermococcus kodakarensis

Cloned(Commentary)

Cloned (Comment)	Organism
gene TK0278 or lysX, genetic organization of the lysine biosynthetic gene cluster, recombinant expression of His6-tagged and of untagged enzyme TkLysX in Escherichia coli strain BL21-CodonPlus (DE3)-RIL	Thermococcus kodakarensis

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant enzyme in complex with product LysW-gamma-AAA, hanging drop vapor diffusion method, method screening and subsequent optimization, mixing of 7.5 mg/ml enzyme, 2.5 mg/ml TkLysW, 10 mM AMP-PNP, 10 mM alpha-aminoadipate (AAA), or 50 mM glutamate, and 10 mM MgSO4 with crystallization solution 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 stucture determination and analysis at 2.18 A resolution	Thermococcus kodakarensis

Crystallization (Comment)

Organism

purified recombinant enzyme in complex with product LysW-gamma-AAA, hanging drop vapor diffusion method, method screening and subsequent optimization, mixing of 7.5 mg/ml enzyme, 2.5 mg/ml TkLysW, 10 mM AMP-PNP, 10 mM alpha-aminoadipate (AAA), or 50 mM glutamate, and 10 mM MgSO4 with crystallization solution 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 stucture determination and analysis at 2.18 A resolution

Thermococcus kodakarensis

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Mg2+	2 magnesium atoms are located in the active site of the enzyme	Thermococcus kodakarensis
phosphate	1 phosphate is located in the active site of the enzyme	Thermococcus kodakarensis

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.
L-lysyl-tRNALys + phosphatidylglycerol	Thermococcus kodakarensis	-	tRNALys + 3-O-L-lysyl-1-O-phosphatidylglycerol	-	?
L-lysyl-tRNALys + phosphatidylglycerol	Thermococcus kodakarensis JCM 12380	-	tRNALys + 3-O-L-lysyl-1-O-phosphatidylglycerol	-	?
L-lysyl-tRNALys + phosphatidylglycerol	Thermococcus kodakarensis ATCC BAA-918	-	tRNALys + 3-O-L-lysyl-1-O-phosphatidylglycerol	-	?
additional information	Thermococcus kodakarensis	enzyme LysX from Thermococcus kodakarensis catalyzes the conjugation of enzyme LysW with either alpha-aminoadipate (AAA) or glutamate	?	-	-
additional information	Thermococcus kodakarensis JCM 12380	enzyme LysX from Thermococcus kodakarensis catalyzes the conjugation of enzyme LysW with either alpha-aminoadipate (AAA) or glutamate	?	-	-
additional information	Thermococcus kodakarensis ATCC BAA-918	enzyme LysX from Thermococcus kodakarensis catalyzes the conjugation of enzyme LysW with either alpha-aminoadipate (AAA) or glutamate	?	-	-

Organism

Organism	UniProt	Comment	Textmining
Thermococcus kodakarensis	Q5JFW0	i.e. Pyrococcus kodakaraensis strain KOD1	-
Thermococcus kodakarensis ATCC BAA-918	Q5JFW0	i.e. Pyrococcus kodakaraensis strain KOD1	-
Thermococcus kodakarensis JCM 12380	Q5JFW0	i.e. Pyrococcus kodakaraensis strain KOD1	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant His6-tagged and of untagged enzyme TkLysX from Escherichia coli strain BL21-CodonPlus (DE3)-RIL, recombinant His6-tagged enzyme by nickel affinity chromatography, recombinant untagged enzyme by ammonium sulfate fractionation, hydrophobic interaction chromatography, ultrafiltration, anion exchange chromatography, gel filtration, and ultrafiltration	Thermococcus kodakarensis

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
L-lysyl-tRNALys + phosphatidylglycerol	-	Thermococcus kodakarensis	tRNALys + 3-O-L-lysyl-1-O-phosphatidylglycerol	-	?
L-lysyl-tRNALys + phosphatidylglycerol	-	Thermococcus kodakarensis JCM 12380	tRNALys + 3-O-L-lysyl-1-O-phosphatidylglycerol	-	?
L-lysyl-tRNALys + phosphatidylglycerol	-	Thermococcus kodakarensis ATCC BAA-918	tRNALys + 3-O-L-lysyl-1-O-phosphatidylglycerol	-	?
additional information	enzyme LysX from Thermococcus kodakarensis catalyzes the conjugation of enzyme LysW with either alpha-aminoadipate (AAA) or glutamate	Thermococcus kodakarensis	?	-	-
additional information	the enzyme LysX also forms LysW-gamma-(alpha-aminoadipate) (LysW-gamma-AAA) from alpha-amino adipate (AAA) and glutamate as substrates and ligates both compounds with TK0279, the amino group carrier protein LysW in this microorganism, in an ATP-dependent manner. LysX, forming an isopeptide bond between the alpha-amino group of AAA and gamma-carboxyl group of the glutamate residue at the C terminus of LysW	Thermococcus kodakarensis	?	-	-
additional information	enzyme LysX from Thermococcus kodakarensis catalyzes the conjugation of enzyme LysW with either alpha-aminoadipate (AAA) or glutamate	Thermococcus kodakarensis JCM 12380	?	-	-
additional information	the enzyme LysX also forms LysW-gamma-(alpha-aminoadipate) (LysW-gamma-AAA) from alpha-amino adipate (AAA) and glutamate as substrates and ligates both compounds with TK0279, the amino group carrier protein LysW in this microorganism, in an ATP-dependent manner. LysX, forming an isopeptide bond between the alpha-amino group of AAA and gamma-carboxyl group of the glutamate residue at the C terminus of LysW	Thermococcus kodakarensis JCM 12380	?	-	-
additional information	enzyme LysX from Thermococcus kodakarensis catalyzes the conjugation of enzyme LysW with either alpha-aminoadipate (AAA) or glutamate	Thermococcus kodakarensis ATCC BAA-918	?	-	-
additional information	the enzyme LysX also forms LysW-gamma-(alpha-aminoadipate) (LysW-gamma-AAA) from alpha-amino adipate (AAA) and glutamate as substrates and ligates both compounds with TK0279, the amino group carrier protein LysW in this microorganism, in an ATP-dependent manner. LysX, forming an isopeptide bond between the alpha-amino group of AAA and gamma-carboxyl group of the glutamate residue at the C terminus of LysW	Thermococcus kodakarensis ATCC BAA-918	?	-	-

Subunits

Subunits	Comment	Organism
tetramer	crystal structure analysis	Thermococcus kodakarensis

Synonyms

Synonyms	Comment	Organism
LysX	-	Thermococcus kodakarensis
RimK-related lysine biosynthesis protein	UniProt	Thermococcus kodakarensis
Tk0278	-	Thermococcus kodakarensis
TkLysX	-	Thermococcus kodakarensis

General Information

General Information	Comment	Organism
evolution	the enzyme belongs to the LysX family of enzymes. Analysis of the mechanism for substrate recognition and its relationship with molecular evolution among LysX family proteins, which have different substrate specificities, overview. Substrate recognition mechanism in LysX family proteins. The lysX homologues in the lysine biosynthetic gene cluster encode enzymes that possess the signature motif (Tyr175, Ile185, Thr196, and Asn250-Ala251) for the LysX/ArgX bifunctional enzyme	Thermococcus kodakarensis
metabolism	the enzyme LysX is involved in the lysine biosynthesis pathway. The conversion from alpha-aminoadipate (AAA) to lysine, is accomplished by five enzymes: LysX, LysZ, LysY, LysJ, and LysK, using the amino group carrier protein LysW. The alpha-amino group of AAA is modified with LysW by LysX, forming an isopeptide bond between the alpha-amino group of AAA and gamma-carboxyl group of the glutamate residue at the C terminus of LysW. LysW-gamma-AAA thus synthesized is then 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. LysW contains many acidic amino acid residues for electrostatic interactions with each enzyme, and, thus, functions as an amino group carrier protein for efficient lysine biosynthesis	Thermococcus kodakarensis
additional information	crystal structure analysis revealing the mechanism of substrate recognition of alpha-aminoadipate substrate by LysX and structural basis for the bifunctionality of the LysX family protein from Thermococcus kodakarensis Active site structure and reaction mechanism, structure-function analysis, detailed overview. ADP, a phosphate ion, two magnesium atoms, and the C-terminus of TkLysW-gamma-AAA are located in the active site of TkLysX	Thermococcus kodakarensis