EC Number | Activating Compound | Comment | Organism | Structure |
---|---|---|---|---|
1.5.1.30 | NADH | - |
Escherichia coli | |
1.5.1.30 | NADPH | - |
Escherichia coli |
EC Number | Cloned (Comment) | Organism |
---|---|---|
1.5.1.30 | PCR-amplification, expression in Escherichia coli Bl21 (deltaDE3) | Escherichia coli |
1.14.14.3 | expressed in Escherichia coli from pJHD500, ligated into a pET21b vector. Luciferase subcloned from pZCH2 into a pASKIBA-3c vector with the restriction sites XbaI and XhoI. The resulting luciferase containing a strep-II tag on the C terminus of the beta-subunit (pZCB4) expressed | Vibrio harveyi |
EC Number | Protein Variants | Comment | Organism |
---|---|---|---|
1.5.1.30 | additional information | Fre oxidoreductase deletion strain shows 99% reduced bioluminescence in coupled assay with luciferase | Escherichia coli |
EC Number | KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|---|
1.5.1.30 | 0.0009 | - |
FMN | coupled oxidoreductase-luciferase assay, kcat/Km = 73/(M*min), in 100 mM Na+/K+ phosphate buffer with 100 mM NaCl, pH 7.0, 1 microM Fre oxidoreductase, 10 microM decanal, 10 microM NADPH, 5 microM luciferase | Escherichia coli | |
1.5.1.30 | 0.0013 | - |
riboflavin | coupled oxidoreductase-luciferase assay, kcat/Km = 1257/(M*min), in 100 mM Na+/K+ phosphate buffer with 100 mM NaCl, pH 7.0, 1 microM Fre oxidoreductase, 10 microM decanal, 10 microM NADPH, 5 microM luciferase, riboflavin is favoured by Fre oxidoreductase but a poor substrate for bacterial luciferase | Escherichia coli | |
1.14.14.3 | 0.0009 | - |
FMN | in the presence of different flavin concentrations, 0.001 mM Fre oxidoreductase, 10 mM decanal, and 0.01 mM NADPH and 0.005 mM luciferase | Vibrio harveyi | |
1.14.14.3 | 0.0013 | - |
riboflavin | in the presence of different flavin concentrations, 0.001 mM Fre oxidoreductase, 10 mM decanal, and 0.01 mM NADPH and 0.005 mM luciferase | Vibrio harveyi |
EC Number | Metals/Ions | Comment | Organism | Structure |
---|---|---|---|---|
1.14.14.3 | K+ | - |
Vibrio harveyi | |
1.14.14.3 | Na+ | - |
Vibrio harveyi |
EC Number | Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|---|
1.5.1.30 | 26000 | - |
SDS-PAGE | Escherichia coli |
EC Number | Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|---|
1.5.1.30 | FMN + NADH + H+ | Escherichia coli | - |
FMNH2 + NAD+ | - |
? | |
1.5.1.30 | FMN + NADPH | Escherichia coli | - |
FMNH2 + NADP+ + H+ | - |
? |
EC Number | Organism | UniProt | Comment | Textmining |
---|---|---|---|---|
1.5.1.30 | Escherichia coli | - |
- |
- |
1.14.14.3 | Vibrio harveyi | - |
- |
- |
EC Number | Purification (Comment) | Organism |
---|---|---|
1.5.1.30 | crude lysate assay with briefly centrifuged cells, resuspended in B-PER bacterial protein extraction reagent, recombinant pZCFRE1 expression product is clarified by centrifugation and applied to a custom nickel affinity column, dialyzed into buffer containing 100 mM Na+/K+ phosphate and 100 mM NaCl, pH 7.0 | Escherichia coli |
1.14.14.3 | nickel affinity column | Vibrio harveyi |
EC Number | Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|---|
1.5.1.30 | FMN + NADH + H+ | - |
Escherichia coli | FMNH2 + NAD+ | - |
? | |
1.5.1.30 | FMN + NADPH | - |
Escherichia coli | FMNH2 + NADP+ + H+ | - |
? | |
1.5.1.30 | FMN + NADPH + H+ | - |
Escherichia coli | FMNH2 + NADP+ | - |
? | |
1.5.1.30 | riboflavin + NADPH + H+ | - |
Escherichia coli | reduced riboflavin + NADP+ | - |
? | |
1.14.14.3 | aldehyde + FMNH2 + O2 | - |
Vibrio harveyi | ? | - |
? | |
1.14.14.3 | decanal + FMNH + O2 | - |
Vibrio harveyi | decanoic acid + FMN + H2O + light | - |
? | |
1.14.14.3 | decanal + riboflavin + O2 | riboflavin is a very poor substrate for bacterial luciferase | Vibrio harveyi | ? | - |
? |
EC Number | Subunits | Comment | Organism |
---|---|---|---|
1.5.1.30 | monomer | 1 * 26000 | Escherichia coli |
EC Number | Synonyms | Comment | Organism |
---|---|---|---|
1.5.1.30 | flavin oxidoreductase | - |
Escherichia coli |
1.5.1.30 | flavin reductase | - |
Escherichia coli |
1.5.1.30 | Fre oxidoreductase | - |
Escherichia coli |
1.14.14.3 | bacterial luciferase | - |
Vibrio harveyi |
EC Number | Cofactor | Comment | Organism | Structure |
---|---|---|---|---|
1.14.14.3 | FMNH | - |
Vibrio harveyi | |
1.14.14.3 | FMNH2 | - |
Vibrio harveyi |
EC Number | General Information | Comment | Organism |
---|---|---|---|
1.5.1.30 | physiological function | provides reducing equivalents required to drive the luciferase genes in Escherichia coli but is not coupled to luciferase | Escherichia coli |
1.14.14.3 | physiological function | bacterial luciferase requires an exogenous source of reduced flavin mononucleotide for bioluminescence activity. There is no stable complex formation between luciferase and oxidoreductases Fre/NfsA from Escherichia coli or Frp from Vibrio harveyi, which are believed to provide reduced flavin for luciferase activity. No difference in the levels of luciferase expression in either the DELTAfre or DELTANsfA strains | Vibrio harveyi |
EC Number | kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|---|
1.5.1.30 | 0.0012 | - |
FMN | coupled oxidoreductase-luciferase assay, in 100 mM Na+/K+ phosphate buffer with 100 mM NaCl, pH 7.0, 1 microM Fre oxidoreductase, 10 microM decanal, 10 microM NADPH, 5 microM luciferase | Escherichia coli | |
1.5.1.30 | 0.021 | - |
riboflavin | coupled oxidoreductase-luciferase assay, in 100 mM Na+/K+ phosphate buffer with 100 mM NaCl, pH 7.0, 1 microM Fre oxidoreductase, 10 microM decanal, 10 microM NADPH, 5 microM luciferase, riboflavin is favoured by Fre oxidoreductase but a poor substrate for bacterial luciferase | Escherichia coli |