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Literature summary extracted from

  • Luttgeharm, K.D.; Cahoon, E.B.; Markham, J.E.
    Substrate specificity, kinetic properties and inhibition by fumonisin B1 of ceramide synthase isoforms from Arabidopsis (2016), Biochem. J., 473, 593-603 .
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.3.1.24 gene LAG2, recombinant expression of codon-optimized LAG2 as N-terminally FLAG-tagged enzyme in Saccharomyces cerevisiae membranes Arabidopsis thaliana
2.3.1.297 gene LAG1, recombinant expression of codon-optimized LAG1 as N-terminally FLAG-tagged enzyme in Saccharomyces cerevisiae membranes Arabidopsis thaliana
2.3.1.297 gene LAG3, recombinant expression of codon-optimized LAG3 as N-terminally FLAG-tagged enzyme in Saccharomyces cerevisiae membranes Arabidopsis thaliana

Protein Variants

EC Number Protein Variants Comment Organism
2.3.1.24 additional information complementation of the growth defect of DELTAlag1/DELTAlac1 yeast deletion mutant by recombinant GST-/FLAG-tagged LOH2. Complementation with LOH2 leads to the accumulation of C16-containing inositolphosphoceramides and ceramides indicating this isoform's preference for C16 fatty acids. Microsomes isolated from LOH2 overexpressing plants showed high levels of C16-ceramide synthase activity compared to microsomes from wild-type plants, indicating that LOH2 overexpression results in accumulation of functional enzyme Arabidopsis thaliana
2.3.1.297 additional information complementation of the growth defect of DELTAlag1/DELTAlac1 yeast deletion mutant by recombinant GST-/FLAG-tagged LOH1. Upon isolation of a microsomal membrane fraction from yeast expressing the construct, LOH1 shows significant activity by an in vitro ceramide synthase assay Arabidopsis thaliana
2.3.1.297 additional information complementation of the growth defect of DELTAlag1/DELTAlac1 yeast deletion mutant by recombinant GST-/FLAG-tagged LOH3. Upon isolation of a microsomal membrane fraction from yeast expressing the construct, LOH3 shows significant activity by an in vitro ceramide synthase assay Arabidopsis thaliana

Inhibitors

EC Number Inhibitors Comment Organism Structure
2.3.1.24 fumonisin B1 mixed mode of inhibition with respect to the long-chain base Arabidopsis thaliana
2.3.1.297 fumonisin B1 mixed mode of inhibition with respect to the laong-chain base; mixed mode of inhibition with respect to the long-chain base Arabidopsis thaliana
2.3.1.297 additional information LOH1 is inhibited by most or all divalent cations tested; LOH3 is inhibited by most or all divalent cations tested Arabidopsis thaliana

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.3.1.24 additional information
-
 additional information Michaelis-Menten kinetics Arabidopsis thaliana
2.3.1.24 0.023
-
 sphinganine pH 7.5, 30°C, LOH2 Arabidopsis thaliana
2.3.1.297 additional information
-
 additional information Michaelis-Menten kinetics Arabidopsis thaliana
2.3.1.297 0.0069
-
 phytosphingosine pH 7.5, 30°C, LOH1 Arabidopsis thaliana
2.3.1.297 0.023
-
 phytosphingosine pH 7.5, 30°C, LOH3 Arabidopsis thaliana

Localization

EC Number Localization Comment Organism GeneOntology No. Textmining
2.3.1.24 microsome
-
 Arabidopsis thaliana
-
-
2.3.1.297 microsome
-
 Arabidopsis thaliana
-
-

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.3.1.24 Ca2+ activates Arabidopsis thaliana
2.3.1.24 Mg2+ activates at 2 mM Arabidopsis thaliana
2.3.1.24 Mn2+ activates Arabidopsis thaliana
2.3.1.24 additional information poor effect observed by 0.001 mM of Cu2+, Zn2+, or Co2+ Arabidopsis thaliana
2.3.1.297 additional information LOH1 is inhibited by most or all divalent cations tested Arabidopsis thaliana
2.3.1.297 additional information LOH3 is inhibited by most or all divalent cations tested Arabidopsis thaliana

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.3.1.24 palmitoyl-CoA + sphinganine Arabidopsis thaliana
-
 CoA + N-palmitoylsphinganine
-
 ?
2.3.1.24 palmitoyl-CoA + sphingosine Arabidopsis thaliana
-
 CoA + N-palmitoylsphingosine
-
 ?
2.3.1.297 hexacosanoyl-CoA + phytosphingosine Arabidopsis thaliana
-
 CoA + N-hexacosanoylphytosphingosine
-
 ?
2.3.1.297 lignoceroyl-CoA + phytosphingosine Arabidopsis thaliana
-
 N-lignoceroylphytosphinganine + CoA
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.3.1.24 Arabidopsis thaliana Q9LJK3
-
-
2.3.1.297 Arabidopsis thaliana Q6NQI8
-
-
2.3.1.297 Arabidopsis thaliana Q9LDF2
-
-

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.3.1.24 additional information specificity of three different isoforms of ceramide synthase, denoted LOH1, 2 and 3, for a range of long-chain base (LCB) and acyl-CoA substrates. The contribution of each of these isoforms to the synthesis of ceramide is investigated by in vitro ceramide synthase assays, overview. The plant LCB phytosphingosine is efficiently used by the LOH1 and 3 isoforms, with LOH1 having the lowest Km for the LCB substrate of the three isoforms. In contrast, sphinganine is used efficiently only by the LOH2 isoform. Acyl-CoA specificity is also distinguished between the three isoforms with LOH2 being almost completely specific for palmitoyl-CoA, while the LOH1 isoform shows greatest activity with lignoceroyl- and hexacosanoyl-CoA. Unsaturated acyl-CoAs are not used efficiently by any isoform while unsaturated LCB substrates are preferred by LOH2 and 3. LOH2 shows poor activity with phytosphingosine. LOH2 is unable to use any acyl-CoA substrates greater than C18 and shows very strong preference for C16 acyl-CoA. Recombinant LOH2 shows maximum activity with the d18:1(4E) LCB about 6 times that with the d18:0 LCB, eventhough d18:1(4E) is not observed in Arabidopsis leaves. Rather d18:2(4E/8Z) or d18:2(4E/8E) are found in selected tissues such as pollen. LOH2 shows enhanced activity towards DELTA4 unsaturated LCB substrates Arabidopsis thaliana ?
-
 ?
2.3.1.24 palmitoyl-CoA + sphinganine
-
 Arabidopsis thaliana CoA + N-palmitoylsphinganine
-
 ?
2.3.1.24 palmitoyl-CoA + sphingosine
-
 Arabidopsis thaliana CoA + N-palmitoylsphingosine
-
 ?
2.3.1.297 acyl-CoA + sphinganine very low activity with sphinganine Arabidopsis thaliana N-acylsphinganine + CoA
-
 ?
2.3.1.297 hexacosanoyl-CoA + phytosphingosine
-
 Arabidopsis thaliana CoA + N-hexacosanoylphytosphingosine
-
 ?
2.3.1.297 lignoceroyl-CoA + phytosphingosine
-
 Arabidopsis thaliana N-lignoceroylphytosphinganine + CoA
-
 ?
2.3.1.297 lignoceroyl-CoA + phytosphingosine high activity Arabidopsis thaliana N-lignoceroylphytosphinganine + CoA
-
 ?
2.3.1.297 additional information specificity of three different isoforms of ceramide synthase, denoted LOH1, 2 and 3, for a range of long-chain base (LCB) and acyl-CoA substrates. The contribution of each of these isoforms to the synthesis of ceramide is investigated by in vitro ceramide synthase assays, overview. The plant LCB phytosphingosine is efficiently used by the LOH1 and 3 isoforms, with LOH1 having the lowest Km for the LCB substrate of the three isoforms. In contrast, sphinganine is used efficiently only by the LOH2 isoform. Acyl-CoA specificity is also distinguished between the three isoforms with LOH2 being almost completely specific for palmitoyl-CoA, while the LOH1 isoform shows greatest activity with lignoceroyl- and hexacosanoyl-CoA. Unsaturated acyl-CoAs are not used efficiently by any isoform while unsaturated LCB substrates are preferred by LOH2 and 3. LOH3 has at least twice as much activity with t18:1 substrates as LOH1. Both LOH1 and LOH3 demonstrate a strong preference for very long chain acyl-CoAs (>C18) although LOH1 has the greatest activity toward 24 and 26 carbon acyl-CoAs, while LOH3 shows little preference for acyl-CoAs between 20 and 26 carbons in length Arabidopsis thaliana ?
-
 ?

Synonyms

EC Number Synonyms Comment Organism
2.3.1.24 ceramide synthase
-
 Arabidopsis thaliana
2.3.1.24 LAG1 longevity assurance homolog 2
-
 Arabidopsis thaliana
2.3.1.24 LAG12
-
 Arabidopsis thaliana
2.3.1.24 LOH2
-
 Arabidopsis thaliana
2.3.1.297 ceramide synthase
-
 Arabidopsis thaliana
2.3.1.297 LAG1 longevity assurance homolog 1
-
 Arabidopsis thaliana
2.3.1.297 LAG1 longevity assurance homolog 3
-
 Arabidopsis thaliana
2.3.1.297 LAG11
-
 Arabidopsis thaliana
2.3.1.297 LAG13
-
 Arabidopsis thaliana
2.3.1.297 LOH1
-
 Arabidopsis thaliana
2.3.1.297 LOH3
-
 Arabidopsis thaliana

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.3.1.24 30
-
 assay at Arabidopsis thaliana
2.3.1.297 30
-
 assay at Arabidopsis thaliana

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.3.1.24 7.5
-
 assay at Arabidopsis thaliana
2.3.1.297 7.5
-
 assay at Arabidopsis thaliana

Ki Value [mM]

EC Number Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
2.3.1.24 0.00097
-
 fumonisin B1 pH 7.5, 30°C, LOH2 Arabidopsis thaliana
2.3.1.297 0.000003
-
 fumonisin B1 pH 7.5, 30°C, LOH1 Arabidopsis thaliana
2.3.1.297 0.000784
-
 fumonisin B1 pH 7.5, 30°C, LOH3 Arabidopsis thaliana

General Information

EC Number General Information Comment Organism
2.3.1.24 physiological function distinct roles and modes of regulation for each of the ceramide synthases in Arabidopsis thaliana sphingolipid metabolism. Ceramides are organizing components of sphingolipids in the eukaryotic cell. Three ceramide synthase isoforms are found in Arabidopsis thaliana each with specific substrate preferences and sensitivity to inhibitors and activators, overview. Isozyme LOH2 is required for the synthesis of ceramides with dihydroxy-long-chain bases and C16 fatty acids Arabidopsis thaliana
2.3.1.297 physiological function distinct roles and modes of regulation for each of the ceramide synthases in Arabidopsis thaliana sphingolipid metabolism. Ceramides are organizing components of sphingolipids in the eukaryotic cell. Three ceramide synthase isoforms are found in Arabidopsis thaliana each with specific substrate preferences and sensitivity to inhibitors and activators, overview. Isozymes LOH1 and LOH3 are responsible for the synthesis of ceramides with trihydroxy long-chain bases and very long chain fatty acids (VLCFA) Arabidopsis thaliana