Literature summary for 1.1.1.27 extracted from

Kolappan, S.; Shen, D.L.; Mosi, R.; Sun, J.; McEachern, E.J.; Vocadlo, D.J.; Craig, L.
Structures of lactate dehydrogenase A (LDHA) in apo, ternary and inhibitor-bound forms (2015), Acta Crystallogr. Sect. D, 71, 185-195.

Cloned(Commentary)

Cloned (Comment)	Organism
recombinant expression of C-terminally His6-tagged enzyme from pET-29b-LDHA vector in Escherichia coli strain BL21(lambdaDE3)	Homo sapiens

Crystallization (Commentary)

Crystallization (Comment)	Organism
lactate dehydrogenase A in apo form, in ternary complex with oxalate and kanamycin, and in inhibitor-bound form, hanging drop vapour diffusion method, apo-LDHA crystals are grown by mixing of 0.002 ml of 25 mg/ml enzyme protein with 0.002 ml reservoir solution consisting of 100 mM Bis-Tris propane, pH 7.0, 20% v/v PEG 400, and 100 mM LiCl. LDHA-NADH crystals are grown by first incubating 25 mg/ml enzyme protein with 3 mM NADH for 2 h at 4°C and then setting up 0.004 ml drops with a 2:1:1 volume ratio of LDHA-NADH, reservoir solution, containing 18% w/v PEG 3350, and 50 mM HEPES, pH 6.8, and another reagent containing 0.16% w/v L-citrulline, 0.16% w/v L-ornithine hydrochloride, 0.16% w/v urea, 0.16% w/v oxalic acid, 0.16% w/v kanamycin monosulfate, and 0.16% w/v L-arginine in 0.02 M HEPES sodium, pH 6.8, 20°C. Apo and NADH-bound LDHA crystals appear after three weeks. LDHA-inhibitor complex crystals are obtained by adding 0.010 ml soaking solution containing 20 mM inhibitor, 100 mM HEPES, pH 7.5, 50 mM LiCl, 100 mM bis-tris propane pH 7.0, 20% v/v DMSO, and 25% PEG 8000, to 0.002 ml hanging drops containing apo LDHA crystals and allowing the crystals to sit at room temperature for 24 h, X-ray diffraction structure determination and analysis at 2.1-3.2 A resolution, molecular replacement method using model structure PDB ID1i10	Homo sapiens

Crystallization (Comment)

Organism

lactate dehydrogenase A in apo form, in ternary complex with oxalate and kanamycin, and in inhibitor-bound form, hanging drop vapour diffusion method, apo-LDHA crystals are grown by mixing of 0.002 ml of 25 mg/ml enzyme protein with 0.002 ml reservoir solution consisting of 100 mM Bis-Tris propane, pH 7.0, 20% v/v PEG 400, and 100 mM LiCl. LDHA-NADH crystals are grown by first incubating 25 mg/ml enzyme protein with 3 mM NADH for 2 h at 4°C and then setting up 0.004 ml drops with a 2:1:1 volume ratio of LDHA-NADH, reservoir solution, containing 18% w/v PEG 3350, and 50 mM HEPES, pH 6.8, and another reagent containing 0.16% w/v L-citrulline, 0.16% w/v L-ornithine hydrochloride, 0.16% w/v urea, 0.16% w/v oxalic acid, 0.16% w/v kanamycin monosulfate, and 0.16% w/v L-arginine in 0.02 M HEPES sodium, pH 6.8, 20°C. Apo and NADH-bound LDHA crystals appear after three weeks. LDHA-inhibitor complex crystals are obtained by adding 0.010 ml soaking solution containing 20 mM inhibitor, 100 mM HEPES, pH 7.5, 50 mM LiCl, 100 mM bis-tris propane pH 7.0, 20% v/v DMSO, and 25% PEG 8000, to 0.002 ml hanging drops containing apo LDHA crystals and allowing the crystals to sit at room temperature for 24 h, X-ray diffraction structure determination and analysis at 2.1-3.2 A resolution, molecular replacement method using model structure PDB ID1i10

Homo sapiens

Inhibitors

Inhibitors	Comment	Organism	Structure
3-([3-carbamoyldimethoxypyrimidin-7-(2,4-dimethoxypyrimidin-5-yl)quinolin-4-yl]amino)benzoic acid	enzyme binding structure, overview	Homo sapiens
3-[7-(2,4-dimethoxypyrimidin-5-yl)-3-sulfamoylquinolin-4-yl]aminobenzoic acid	enzyme binding structure, overview	Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
(S)-lactate + NAD+	Homo sapiens	-	pyruvate + NADH + H+	-	r

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	P00338	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant C-terminally His6-tagged enzyme from Escherichia coli strain BL21(lambdaDE3) by centrfugation at 40000 g, nickel affinity chromatography, dialysis and ultrafiltration, followed by gel filtration, to over 95% purity	Homo sapiens

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
(S)-lactate + NAD+	-	Homo sapiens	pyruvate + NADH + H+	-	r

Subunits

Subunits	Comment	Organism
More	LDHA ternary structures, overview	Homo sapiens
tetramer	crystal structure analysis	Homo sapiens

Synonyms

Synonyms	Comment	Organism
lactate dehydrogenase A	-	Homo sapiens
LdhA	-	Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at, pyruvate reduction	Homo sapiens

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at, pyruvate reduction	Homo sapiens

Cofactor

Cofactor	Comment	Organism	Structure
NAD+	-	Homo sapiens
NADH	-	Homo sapiens

Ki Value [mM]

Ki Value [mM]	Ki Value maximum [mM]	Inhibitor	Comment	Organism	Structure
0.00059	-	3-[7-(2,4-dimethoxypyrimidin-5-yl)-3-sulfamoylquinolin-4-yl]aminobenzoic acid	pH 7.5, 37°C, recombinant His-tagged enzyme, pyruvate reduction, competitive versus NADH	Homo sapiens
0.00252	-	3-[7-(2,4-dimethoxypyrimidin-5-yl)-3-sulfamoylquinolin-4-yl]aminobenzoic acid	pH 7.5, 37°C, recombinant His-tagged enzyme, pyruvate reduction, noncompetitive versus pyruvate	Homo sapiens

IC50 Value

IC50 Value	IC50 Value Maximum	Comment	Organism	Inhibitor
0.000039	-	pH 7.5, 37°C, recombinant His-tagged enzyme, L-lactate oxidation	Homo sapiens	3-[7-(2,4-dimethoxypyrimidin-5-yl)-3-sulfamoylquinolin-4-yl]aminobenzoic acid
0.00051	-	pH 7.5, 37°C, recombinant His-tagged enzyme, L-lactate oxidation	Homo sapiens	3-([3-carbamoyldimethoxypyrimidin-7-(2,4-dimethoxypyrimidin-5-yl)quinolin-4-yl]amino)benzoic acid
0.0022	-	pH 7.5, 37°C, recombinant His-tagged enzyme, pyruvate reduction	Homo sapiens	3-[7-(2,4-dimethoxypyrimidin-5-yl)-3-sulfamoylquinolin-4-yl]aminobenzoic acid
0.0144	-	pH 7.5, 37°C, recombinant His-tagged enzyme, pyruvate reduction	Homo sapiens	3-([3-carbamoyldimethoxypyrimidin-7-(2,4-dimethoxypyrimidin-5-yl)quinolin-4-yl]amino)benzoic acid