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Literature summary extracted from

  • Wang, Z.; Min, X.; Xiao, S.H.; Johnstone, S.; Romanow, W.; Meininger, D.; Xu, H.; Liu, J.; Dai, J.; An, S.; Thibault, S.; Walker, N.
    Molecular basis of sphingosine kinase 1 substrate recognition and catalysis (2013), Structure, 21, 798-809.
    View publication on PubMed

Application

EC Number Application Comment Organism
2.7.1.91 drug development the enzyme is a potential target for drug discovery Homo sapiens

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.7.1.91 recombinant expression of N-terminally His6-tagged wild-type and mutant enzymes in High Five cells using the baculovirus transfection method Homo sapiens

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
2.7.1.91 purified recombinant truncated enzyme comprising residues 9-364, in apo-form and in complex with inhibitor SKI-II, and with inhibitor SKI-II and ADP, sitting drop vapour diffusion method, mixing of 0.001 ml of 10 mg/ml protein in 50 mM Tris-HCl, pH 8.5, 300 mM NaCl, 0.5 M GdnHCl, 10% glycerol, 0.1% Tween 20, and 10 mM DTT, and for complex crystals with a 3fold excess of inhibitor, and ATP/ADP, with 0.001 ml of well solution containing 1.0-1.4 M ammonium sulfate, 0.3-1.3 M NaCl, and 0.1 M Bis-Tris, pH 6.0, 16°C, X-ray diffraction structure determination and analysis at 2.0-2.3 A resolution, molecular replacement Homo sapiens

Protein Variants

EC Number Protein Variants Comment Organism
2.7.1.91 D81A site-directed mutagenesis, the mutant shows essentially no activity for phosphoryl transfer and ADP formation Homo sapiens
2.7.1.91 D81N site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme Homo sapiens
2.7.1.91 additional information construction of an N- and C-terminally truncated enzyme form comprising residues 9-364 of full-length 368 residues, the mutant is catalytically active Homo sapiens

Inhibitors

EC Number Inhibitors Comment Organism Structure
2.7.1.91 SKI-II highly selective and non ATP-competitive, enzyme binding structure, overview Homo sapiens

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.7.1.91 additional information
-
 additional information Michaelis-Menten kinetics Homo sapiens
2.7.1.91 0.02
-
 ATP recombinant wild-type enzyme, pH and temperature not specified in the publication Homo sapiens
2.7.1.91 0.0255
-
 ATP recombinant mutant D81N enzyme, pH and temperature not specified in the publication Homo sapiens

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.7.1.91 Mg2+ required, binding structure, overview Homo sapiens

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.7.1.91 ATP + sphingosine Homo sapiens
-
 ADP + sphingosine 1-phosphate
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.7.1.91 Homo sapiens Q9NYA1
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.7.1.91 recombinant N-terminally His6-tagged wild-type and mutant enzymes from High Five cells by nickel affinity chromatography, the tag is cleaved by TEV protease, followed by anion and cation exchange chromatographies, and gel filtration Homo sapiens

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.7.1.91 ATP + D-erythro-sphingosine
-
 Homo sapiens ADP + D-erythro-sphingosine 1-phosphate nucleotide binding in the cleft between the N-terminal domain and C-terminal domain ?
2.7.1.91 ATP + sphingosine
-
 Homo sapiens ADP + sphingosine 1-phosphate
-
 ?

Subunits

EC Number Subunits Comment Organism
2.7.1.91 More the SphK1 structure reveals a two-domain architecture in which its catalytic site is located in the cleft between the two domains and a hydrophobic lipid-binding pocket is buried in the C-terminal domain, structure comparisons with Staphylococcus aureus DgkB, PDB ID 2QVL, and Archaeoglobus fulgidus NAD kinase, PDB ID 1ZOZ Homo sapiens

Synonyms

EC Number Synonyms Comment Organism
2.7.1.91 sphingosine kinase 1
-
 Homo sapiens
2.7.1.91 SPHK1
-
 Homo sapiens

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.7.1.91 0.0065
-
 ATP recombinant mutant D81N enzyme, pH and temperature not specified in the publication Homo sapiens
2.7.1.91 0.0427
-
 ATP recombinant wild-type enzyme, pH and temperature not specified in the publication Homo sapiens

Cofactor

EC Number Cofactor Comment Organism Structure
2.7.1.91 ATP
-
 Homo sapiens

General Information

EC Number General Information Comment Organism
2.7.1.91 additional information modeling of the catalytic mechanism, overview. Residue Asp81 plays a critical role in the phosphoryl transfer step, but not in the binding of ATP Homo sapiens
2.7.1.91 physiological function lipid kinase sphingosine kinase 1 catalyzes the conversion of sphingosine to sphingosine-1-phosphate, which plays a role in lymphocyte trafficking, angiogenesis, and response to apoptotic stimuli. As a central enzyme in modulating the sphingosine-1-phosphate levels in cells, the enzyme is an important regulator for diverse cellular functions Homo sapiens