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Literature summary for 1.13.12.6 extracted from

  • Mitani, Y.; Oshima, Y.; Mitsuda, N.; Tomioka, A.; Sukegawa, M.; Fujita, M.; Kaji, H.; Ohmiya, Y.
    Efficient production of glycosylated Cypridina luciferase using plant cells (2017), Protein Expr. Purif., 133, 102-109 .
    View publication on PubMed

Application

Application Comment Organism
analysis Cypridina noctiluca luciferase is utilized for biochemical and molecular biological applications, including bioluminescent enzyme immunoassays, far-red luminescence imaging, and high-throughput reporter assays Cypridina noctiluca
molecular biology Cypridina noctiluca luciferase is utilized for biochemical and molecular biological applications, including bioluminescent enzyme immunoassays, far-red luminescence imaging, and high-throughput reporter assays Cypridina noctiluca

Cloned(Commentary)

Cloned (Comment) Organism
recombinant expression from vector p35SHSPG in Nicotiana benthamiana BY-2 cells, luciferase is efficiently secreted from cells with its native signal peptide, two potential N-glycosylation sites are modified with plant-type oligosaccharide chains. Recombinant expression of the enzyme in Arabidopsis thaliana roots in the cell wall of intact cells and the apoplast of plasmolyzed cells but not in the protoplasm. Cluc contains its own secretion signal peptide, which results in efficient secretion and folding of Cluc Cypridina noctiluca

Protein Variants

Protein Variants Comment Organism
additional information for large scale enzyme production, a simple production procedure is established using plant cell cultures. The plant cell culture tobacco BY-2 efficiently secretes luciferase, which is easily purified using a simple one-step ion-exchange chromatography method. The production yield is 20-30 mg of luciferase per liter of culture medium. The method offers a cost-effective production for Cypridina luciferase Cypridina noctiluca

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten kinetics Cypridina noctiluca
0.00028
-
Cypridina luciferin recombinant enzyme, pH and temperature not specified in the publication Cypridina noctiluca
0.00045
-
Vargula hilgendorfii luciferin recombinant enzyme, recombinant substrate, pH and temperature not specified in the publication Cypridina noctiluca
0.00052
-
Vargula hilgendorfii luciferin recombinant enzyme, native substrate, pH and temperature not specified in the publication Cypridina noctiluca

Localization

Localization Comment Organism GeneOntology No. Textmining
extracellular the enzyme Cluc contains a secretion signal peptide, which results in efficient secretion and folding of Cluc Cypridina noctiluca
-
-

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
Cypridina luciferin + O2 Cypridina noctiluca
-
oxidized Cypridina luciferin + CO2 + hv
-
?

Organism

Organism UniProt Comment Textmining
Cypridina noctiluca Q75R40
-
-

Posttranslational Modification

Posttranslational Modification Comment Organism
glycoprotein glycosylation pattern analysis using LCMS/MS. In the recombinnat enzyme, two potential N-glycosylation sites are modified with plant-type oligosaccharide chains. Partial deglycosylation (trimming) of recombinant Cluc by beta-N-acetylhexosaminidase, HexNAc residues at the non-reducing termini of glycopeptides (Pronase E-digested) are completely eliminated at both glycosylation sites after beta-N-acetylhexosaminidase treatment (at Asn182 and Asn404). The glycan compositions after treatment displays trimannosyl cores (Hex3HexNAc2)+/-Fuc+/-Xyl. Specifically, because the trimannosyl core is fully occupied by Fuc and Xyl on Asn404, only a single signal corresponding to the trimannosyl core +Fuc +Xyl remains after treatment. This glycan-trimmed recombinant protein still exhibits nearly the same level of activity as the unmodified glycoprotein Cypridina noctiluca

Purification (Commentary)

Purification (Comment) Organism
recombinant secreted enzyme from Nicotiana benthamiana BY-2 cell medium by ultrafiltration, buffer exchange gel filtration, anion exchange chromatography, and dialysis Cypridina noctiluca

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
Cypridina luciferin + O2
-
Cypridina noctiluca oxidized Cypridina luciferin + CO2 + hv
-
?
Vargula hilgendorfii luciferin + O2 native or recombinant substrate protein Cypridina noctiluca oxidized Vargula hilgendorfii luciferin + CO2 + hv
-
?

Subunits

Subunits Comment Organism
? x * 61000, recombinnat glycolysated enzyme, SDS-PAGE Cypridina noctiluca

Synonyms

Synonyms Comment Organism
CLuc
-
Cypridina noctiluca
Cypridina luciferase
-
Cypridina noctiluca