Activating Compound | Comment | Organism | Structure |
---|---|---|---|
small subunits of serine palmitoyltransferase | small subunits of SPT, ssSPTa and ssSPTb, increase human SPT activity by 50-100fold when coexpressed with enzyme subunits hLCB1 and hLCBa | Homo sapiens |
Cloned (Comment) | Organism |
---|---|
recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) | Sphingomonas paucimobilis |
recombinant expression of subunit LCB2a mutants in HEK293 cells, recombinant His-tagged enzyme expression in Escherichia coli strain BL21(DE3). Recombinant expression of wild-type hLCB1/hLCB2a subunits in Saccharomyces cerevisiae microsomes in the absence of either small subunit, the enzyme shows very low activity. Coexpression of each of the three subunit hLCB2a mutants, V359M, G382V, and I504F, along with subunit hLCB1 results in low activity, with G382V barely detectable above background | Homo sapiens |
Protein Variants | Comment | Organism |
---|---|---|
C133W | naturally occuring mutation in subunit LCB1 involved in hereditary sensory and autonomic neuropathy type I disease, the mutant shows reduced activity compared tot he wild-type enzyme | Homo sapiens |
C133Y | naturally occuring mutation in subunit LCB1 involved in hereditary sensory and autonomic neuropathy type I disease, the mutant shows reduced activity compared tot he wild-type enzyme | Homo sapiens |
G268V | site-directed mutagenesis, the mutation perturbs the pyridoxal 5'-phosphate cofactor binding and reduces the affinity for both substrates, inactive mutant, the protein is expressed in a completely insoluble form, structure homology modeling of the mutant enzyme using the Sp SPT PLP-L-serine external aldimine structure, PDB ID 2W8J | Sphingomonas paucimobilis |
G382V | naturally occuring mutation in subunit LCB2a involved in hereditary sensory and autonomic neuropathy type I disease, the activity of mutant G382V is barely detectable above background | Homo sapiens |
G385F | site-directed mutagenesis, the mutation perturbs the pyridoxal 5'-phosphate cofactor binding, reduces the affinity for both substrates, decreases the enzyme activity, soluble protein | Sphingomonas paucimobilis |
I504F | naturally occuring mutation in subunit LCB2a involved in hereditary sensory and autonomic neuropathy type I disease, the mutant shows reduced activity compared tot he wild-type enzyme | Homo sapiens |
additional information | an enzyme knockout strain, which completely lacks SPT activity, is not viable unless supplemented with a long chain base or a competent, active SPT complex. Coexpression of each of the three subunit hLCB2a mutants, V359M, G382V, and I504F, along with subunit hLCB1 results in low activity, with G382V barely detectable above background. When small subunit ssSPTb is expressed, heterodimers containing the G382V and I504F mutant hLCB2a subunits are activated to the same extent as wild-type heterodimers, but heterodimers containing the V359Mmutant subunit are less well activated | Homo sapiens |
V246M | site-directed mutagenesis, the mutation perturbs the pyridoxal 5'-phosphate cofactor binding, reduces the affinity for both substrates, decreases the enzyme activity, soluble protein | Sphingomonas paucimobilis |
V359M | naturally occuring mutation in subunit LCB2a involved in hereditary sensory and autonomic neuropathy type I disease, the mutant shows reduced activity compared tot he wild-type enzyme | Homo sapiens |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten kinetics of wild-type and mutant enzymes | Sphingomonas paucimobilis | |
additional information | - |
additional information | Michaelis-Menten kinetics of wild-type and mutant enzymes | Homo sapiens | |
0.0356 | - |
palmitoyl-CoA | recombinant wild-type enzyme, pH 7.5, 25°C | Sphingomonas paucimobilis | |
0.0522 | - |
palmitoyl-CoA | recombinant mutant G385F, pH 7.5, 25°C | Sphingomonas paucimobilis | |
0.128 | - |
palmitoyl-CoA | recombinant mutant V246M, pH 7.5, 25°C | Sphingomonas paucimobilis | |
1.6 | - |
L-serine | recombinant wild-type enzyme, pH 7.5, 25°C | Sphingomonas paucimobilis | |
2.5 | - |
L-serine | recombinant mutant V246M, pH 7.5, 25°C | Sphingomonas paucimobilis | |
3 | - |
L-serine | recombinant mutant G385F, pH 7.5, 25°C | Sphingomonas paucimobilis |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|---|---|---|---|
membrane | membrane-bound enzyme | Homo sapiens | 16020 | - |
microsome | - |
Homo sapiens | - |
- |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
90000 | - |
about, recombinant wild-type enzyme and mutants V246M and G385F, gel filtration | Sphingomonas paucimobilis |
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 | - |
? | |
palmitoyl-CoA + L-serine | Homo sapiens | - |
CoA + 3-dehydro-D-sphinganine + CO2 | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Homo sapiens | O15269 and O15270 | subunits hLCB1 (O15269) and hLCB2a/b (O15270) | - |
Sphingomonas paucimobilis | Q93UV0 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography | Homo sapiens |
recombinant His-tagged wild-type and enzyme and mutants V246M and G385F from Escherichia coli strain BL21(DE3) by nickel affinity chromatography | Sphingomonas paucimobilis |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
palmitoyl-CoA + L-serine = CoA + 3-dehydro-D-sphinganine + CO2 | the reaction proceeds via a key carbanion/quinonoid species and an internal aldimine/PLP-bound form of the enzyme | Sphingomonas paucimobilis | |
palmitoyl-CoA + L-serine = CoA + 3-dehydro-D-sphinganine + CO2 | the reaction proceeds via a key carbanion/quinonoid species and an internal aldimine/PLP-bound form of the enzyme | Homo sapiens |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
palmitoyl-CoA + L-serine | - |
Sphingomonas paucimobilis | CoA + 3-dehydro-D-sphinganine + CO2 | - |
? | |
palmitoyl-CoA + L-serine | - |
Homo sapiens | CoA + 3-dehydro-D-sphinganine + CO2 | - |
? | |
S-(2-oxoheptadecyl)-CoA + L-serine | - |
Homo sapiens | CoA + ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
heterodimer | the membrane-bound heterodimer composed of two subunits hLCB1 and hLCB2a/b | Sphingomonas paucimobilis |
heterodimer | the membrane-bound heterodimer composed of two subunits hLCB1 and hLCB2a/b. The hLCB1 and hLCB2 subunits display relatively high sequence homology to each other, but only the hLCB2a subunit contains key catalytic residues including the lysine that binds pyridoxal 5'-phosphate. Subunit hLCB1 lacks these residues but contains other residues involved in catalysis,the hLCB1/hLCB2a heterodimer has a single active site | Homo sapiens |
Synonyms | Comment | Organism |
---|---|---|
serine palmitoyltransferase | - |
Sphingomonas paucimobilis |
serine palmitoyltransferase | - |
Homo sapiens |
SPT | - |
Sphingomonas paucimobilis |
SPT | - |
Homo sapiens |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
assay at | Sphingomonas paucimobilis |
25 | - |
assay at | Homo sapiens |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.41 | - |
L-serine | recombinant mutant G385F, pH 7.5, 25°C | Sphingomonas paucimobilis | |
0.41 | - |
palmitoyl-CoA | recombinant mutant G385F, pH 7.5, 25°C | Sphingomonas paucimobilis | |
0.44 | - |
L-serine | recombinant mutant V246M, pH 7.5, 25°C | Sphingomonas paucimobilis | |
0.44 | - |
palmitoyl-CoA | recombinant mutant V246M, pH 7.5, 25°C | Sphingomonas paucimobilis | |
1.14 | - |
L-serine | recombinant wild-type enzyme, pH 7.5, 25°C | Sphingomonas paucimobilis | |
1.14 | - |
palmitoyl-CoA | recombinant wild-type enzyme, pH 7.5, 25°C | Sphingomonas paucimobilis |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.5 | - |
assay at | Sphingomonas paucimobilis |
7.5 | - |
assay at | Homo sapiens |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
pyridoxal 5'-phosphate | dependent on | Sphingomonas paucimobilis | |
pyridoxal 5'-phosphate | dependent on, bound by lysine 265 residue on subunit LCB2a | Homo sapiens |
General Information | Comment | Organism |
---|---|---|
malfunction | mutations in both subunits hLCB1 (e.g., C133W and C133Y) and hLCB2a (e.g., V359M, G382V, and I504F) are identified in patients with hereditary sensory and autonomic neuropathy type I (HSAN1), an inherited disorder that affects sensory and autonomic neurons. These mutations result in substrate promiscuity, leading to formation of neurotoxic deoxysphingolipids found in affected individuals. Structure homology modeling to understand the impact of the hLCB2a mutations on the mechanism of the enzyme using the structure data from the Sphingomonas paucimobilis enzyme | Homo sapiens |
metabolism | the enzyme catalyses the first step of de novo sphingolipid biosynthesis | Sphingomonas paucimobilis |
metabolism | the enzyme catalyses the first step of de novo sphingolipid biosynthesis | Homo sapiens |
additional information | structure modeling | Sphingomonas paucimobilis |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.136 | - |
palmitoyl-CoA | recombinant mutant V246M, pH 7.5, 25°C | Sphingomonas paucimobilis | |
0.176 | - |
L-serine | recombinant mutant V246M, pH 7.5, 25°C | Sphingomonas paucimobilis | |
0.712 | - |
L-serine | recombinant wild-type enzyme, pH 7.5, 25°C | Sphingomonas paucimobilis | |
3.437 | - |
L-serine | recombinant mutant G385F, pH 7.5, 25°C | Sphingomonas paucimobilis | |
7.854 | - |
palmitoyl-CoA | recombinant mutant G385F, pH 7.5, 25°C | Sphingomonas paucimobilis | |
32.02 | - |
palmitoyl-CoA | recombinant wild-type enzyme, pH 7.5, 25°C | Sphingomonas paucimobilis |