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Literature summary for 2.3.1.50 extracted from

  • Wadsworth, J.M.; Clarke, D.J.; McMahon, S.A.; Lowther, J.P.; Beattie, A.E.; Langridge-Smith, P.R.; Broughton, H.B.; Dunn, T.M.; Naismith, J.H.; Campopiano, D.J.
    The chemical basis of serine palmitoyltransferase inhibition by myriocin (2013), J. Am. Chem. Soc., 135, 14276-14285.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Sphingomonas paucimobilis

Crystallization (Commentary)

Crystallization (Comment) Organism
determination of the crystal structure of enzyme mutant K265A that diffract to 1.6 A resolution and contain a canonical dimer in the asymmetric unit. Crystallization of the wild-type SPT:PLP-myriocin aldimine complex is not possible, most likely due to aldimine degradation Sphingomonas paucimobilis

Protein Variants

Protein Variants Comment Organism
K265A site-directed mutagenesis, the mutant is unable to bind pyridoxal 5'-phosphate, structure of a SPT K265A:PLP-myriocin external aldimine complex, molecular replacement study Sphingomonas paucimobilis

Inhibitors

Inhibitors Comment Organism Structure
myriocin i.e. (2S,3R,4R,6E)-2-amino-3,4-dihydroxy-2-(hydroxymethyl)-14-oxo-6-eicosenoic acid, or thermozymocidin and ISP-1, is a fungal natural product, kinetics and molecular mechanism of enzyme inhibition, overview. Myriocin is a potent SPT enzyme inhibitor. It initially forms an external aldimine with pyridoxal 5'-phosphate at the active site, the structure of the resulting co-complex explains its nanomolar affinity for the enzyme. The cofactor-inhibitor co-complex, PLP-myriocin aldimine, catalytically degrades via an unexpected retro-aldol-like cleavage mechanism to a C18 aldehyde which in turn acts as a suicide inhibitor of the enzyme by covalent modification of the essential catalytic lysine. This dual mechanism of inhibition rationalizes the extraordinary potency and longevity of myriocin inhibition. Incubations of enzyme SPT with myriocin is consistent with the formation of an initial enzyme-inhibitor complex which is noncovalent in nature and reversible, albeit with a very slow off rate (koff), the PLP-myriocin aldimine as the initial competitive inhibitory species. In contrast, for the enzyme preincubated with myriocin for 16 h, no detectable regain in activity is observed after dialysis either at 3 or 24 h, the second formed covalent adduct acts as an irreversible inhibitor of enzyme SPT Sphingomonas paucimobilis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten kinetics of wild-type and mutant enzyme Sphingomonas paucimobilis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
palmitoyl-CoA + L-serine Sphingomonas paucimobilis
-
CoA + 3-dehydro-D-sphinganine + CO2
-
?

Organism

Organism UniProt Comment Textmining
Sphingomonas paucimobilis Q93UV0
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration Sphingomonas paucimobilis

Reaction

Reaction Comment Organism Reaction ID
palmitoyl-CoA + L-serine = CoA + 3-dehydro-D-sphinganine + CO2 pyridoxal 5'-phosphate dependent reaction mechanism, key active site residues are His159, Asp231, His234, and Lys265 Sphingomonas paucimobilis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
palmitoyl-CoA + L-serine
-
Sphingomonas paucimobilis CoA + 3-dehydro-D-sphinganine + CO2
-
?

Synonyms

Synonyms Comment Organism
serine palmitoyltransferase
-
Sphingomonas paucimobilis
SPT
-
Sphingomonas paucimobilis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Sphingomonas paucimobilis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5 8 assay at Sphingomonas paucimobilis

Cofactor

Cofactor Comment Organism Structure
pyridoxal 5'-phosphate dependent on Sphingomonas paucimobilis

General Information

General Information Comment Organism
metabolism sphingolipids are essential components of cellular membranes formed from the condensation of L-serine and a long-chain acyl thioester. This first step is catalyzed by the pyridoxal 5'-phosphate-dependent enzyme serine palmitoyltransferase Sphingomonas paucimobilis
additional information key active site residues are His159, Asp231, His234, and Lys265 Sphingomonas paucimobilis