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Information on EC 1.13.12.5 - Renilla-type luciferase

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IUBMB Comments
From the soft coral coelenterate Renilla reniformis. The luciferin is bound to a luciferin-binding protein (BP-LH2). The bioluminescent reaction between the luciferin complex, luciferase and oxygen is triggered by calcium ions. In vivo, the excited state luciferin---luciferase complex undergoes the process of nonradiative energy transfer to an accessory protein, Renilla green fluorescent protein, which results in green bioluminescence. In vitro, Renilla luciferase emits blue light in the absence of any green fluorescent protein.
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Word Map
The enzyme appears in viruses and cellular organisms
Reaction Schemes
hide(Overall reactions are displayed. Show all >>)
+
=
excited coelenteramide h monoanion
+
EC NUMBER
COMMENTARY hide
1.13.12.5
-
RECOMMENDED NAME
GeneOntology No.
Renilla-type luciferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
coelenterazine h + O2 = coelenterazine h dioxetanone
show the reaction diagram
(1a)
-
-
-
coelenterazine h + O2 = excited coelenteramide h monoanion + CO2
show the reaction diagram
coelenterazine h dioxetanone = excited coelenteramide h monoanion + CO2
show the reaction diagram
(1b)
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
decarboxylation
Renilla sp.
-
-
oxidation
oxidative decarboxylation
-
-
redox reaction
-
-
-
-
reduction
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
coelenterazine h:oxygen 2-oxidoreductase (decarboxylating)
From the soft coral coelenterate Renilla reniformis. The luciferin is bound to a luciferin-binding protein (BP-LH2). The bioluminescent reaction between the luciferin complex, luciferase and oxygen is triggered by calcium ions. In vivo, the excited state luciferin---luciferase complex undergoes the process of nonradiative energy transfer to an accessory protein, Renilla green fluorescent protein, which results in green bioluminescence. In vitro, Renilla luciferase emits blue light in the absence of any green fluorescent protein.
CAS REGISTRY NUMBER
COMMENTARY hide
346421-46-3
Gaussia luciferase
61869-41-8
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
Renilla sp.
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
-
a collapse of alpha/beta hydrolase fold domain may trigger the irreversible inactivation of the enzyme at higher temperatures. In contrast to wild-type SRLuc8, the alpha-helices in the alpha/beta hydrolase fold domain of engineered C-SRLuc8 have lower perturbations and do not collapse, while some cap domain residues have more perturbations
physiological function
-
Renilla luciferase is an enzyme that is responsible for the bioluminescence of the soft coral Renilla
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
12-benzyl-8-hydroxy-2-(4-hydroxybenzyl)-5,11-dihydrobenzo[f]imidazo[1,2-a]quinoxalin-3(6H)-one + O2
oxidized 12-benzyl-8-hydroxy-2-(4-hydroxybenzyl)-5,11-dihydrobenzo[f]imidazo[1,2-a]quinoxalin-3(6H)-one + CO2 + hnu
show the reaction diagram
-
-
-
-
?
2,8-dibenzyl-6-(4-hydroxyphenyl)imidazo[1,2-a]pyrazin-3(7H)-one + O2
oxidized 2,8-dibenzyl-6-(4-hydroxyphenyl)imidazo[1,2-a]pyrazin-3(7H)-one + CO2 + hn
show the reaction diagram
-
-
-
-
?
2-benzyl-8-benzyl-6-(2-fluorophenylethynyl)imidazo[1,2-a]pyrazin-3(7H)-one + O2
? + CO2 + hv
show the reaction diagram
-
-
-
-
?
2-benzyl-8-benzyl-6-(3-fluorophenylethynyl)imidazo[1,2-a]pyrazin-3(7H)-one + O2
? + CO2 + hv
show the reaction diagram
-
-
-
-
?
2-benzyl-8-benzyl-6-(3-hydroxyphenylethynyl)imidazo[1,2-a]pyrazin-3(7H)-one + O2
? + CO2 + hv
show the reaction diagram
-
-
-
-
?
2-benzyl-8-benzyl-6-(3-methylphenylethynyl)imidazo[1,2-a]pyrazin-3(7H)-one + O2
? + CO2 + hv
show the reaction diagram
-
-
-
-
?
2-benzyl-8-benzyl-6-(4-fluorophenylethynyl)imidazo[1,2-a]pyrazin-3(7H)-one + O2
? + CO2 + hv
show the reaction diagram
-
-
-
-
?
2-benzyl-8-benzyl-6-(phenylethynyl)imidazo[1,2-a]pyrazin-3(7H)-one + O2
? + CO2 + hv
show the reaction diagram
-
-
-
-
?
2-benzyl-8-benzyl-6-[(1-fluoroethyl)-1,2,3-triazol-4]imidazo[1,2-a]pyrazin-3(7H)-one + O2
? + CO2 + hv
show the reaction diagram
-
-
-
-
?
2-benzyl-8-benzyl-6-[(1-hydroxyethyl)-1,2,3-triazol-4]imidazo[1,2-a]pyrazin-3(7H)-one + O2
? + CO2 + hv
show the reaction diagram
-
-
-
-
?
2-benzyl-8-benzyl-6-[(1-hydroxypropyl)-1,2,3-triazol-4]imidazo[1,2-a]pyrazin-3(7H)-one + O2
? + CO2 + hv
show the reaction diagram
-
-
-
-
?
2-hydroperoxycoelenterazine + O2
?
show the reaction diagram
-
-
-
?
3iso-coelenterazine + O2
?
show the reaction diagram
3me-coelenterazine + O2
?
show the reaction diagram
3meo-coelenterazine + O2
?
show the reaction diagram
8-benzyl-2-(4-fluorobenzyl)-6-(4-hydroxyphenyl)imidazo[1,2-a]pyrazin-3(7H)-one + O2
oxidized 8-benzyl-2-(4-fluorobenzyl)-6-(4-hydroxyphenyl)imidazo[1,2-a]pyrazin-3(7H)-one + CO2 + hnu
show the reaction diagram
-
-
-
-
?
8-benzyl-2-(4-hydroxybenzyl)-6-(4-hydroxyphenyl)imidazo[1,2-a]pyrazin-3(7H)-one + O2
oxidized 8-benzyl-2-(4-hydroxybenzyl)-6-(4-hydroxyphenyl)imidazo[1,2-a]pyrazin-3(7H)-one + CO2 + hn
show the reaction diagram
-
-
-
-
?
alphameh-coelenterazine + O2
?
show the reaction diagram
b-coelenterazine + O2
?
show the reaction diagram
Renilla sp.
-
-
-
-
?
beetle D-luciferin
?
show the reaction diagram
-
450 microM
-
-
?
benzylluciferin + O2
oxidized benzylluciferin + CO2 + hv
show the reaction diagram
-
-
-
-
?
benzylluciferin methyl ether + O2
oxidized benzylluciferin methyl ether + CO2 + hv
show the reaction diagram
-
-
-
-
?
bis-coelenterazine + O2
hnu + ?
show the reaction diagram
-
assay at pH 7.6, about 100fold reduced relative luminescence
-
-
?
cf3-coelenterazine + O2
?
show the reaction diagram
coelenterate-type luciferin + O2
oxidized coelenterate-type luciferin + CO2 + hv
show the reaction diagram
-
-
-
-
?
coelenterazine + ?
?
show the reaction diagram
-
20 microM
-
-
?
coelenterazine + O2
?
show the reaction diagram
coelenterazine + O2
coelenteramide + CO2 + hnu
show the reaction diagram
coelenterazine + O2
coelenteramide + CO2 + hv
show the reaction diagram
coelenterazine + O2
oxidized coelenterazine + CO2 + hn
show the reaction diagram
-
-
-
-
?
coelenterazine h + O2
coelenteramide h + CO2 + hv
show the reaction diagram
-
-
-
-
?
coelenterazine-H + O2
?
show the reaction diagram
coelenterazine-h + O2
coelenteramide h + CO2 + hv
show the reaction diagram
-
substrate binding structure
-
-
?
coelenterazine-v + O2
coelenteramide-v + CO2 + hnu
show the reaction diagram
-
increase of substrate coelenterazine stability by ligating it to Ca2+-triggered coelenterazine-binding protein, CBP, from Renilla muelleri, which apparently functions in the organism for stabilizing and protecting coelenterazine from oxidation. The apo-CBP binds coelenterazine-v very rapidly from Ca2+ free solution, similar to that as the native coelenterazine. At low concentrations, coelenterazine-v bound within CBP generates a brighter bioluminescence signal than would free coelenterazine, thereby increasing the assay sensitivity, overview
orange bioluminescence
-
?
coelentrazine + O2
?
show the reaction diagram
-
-
-
-
?
cp-coelenterazine + O2
hnu + ?
show the reaction diagram
-
assay at pH 7.6, about 100fold reduced relative luminescence
-
-
?
D-luciferin + O2
?
show the reaction diagram
D-luciferin + O2 + ATP
oxidized D-luciferin + CO2 + H2O + AMP + diphosphate + hv
show the reaction diagram
e-coelenterazine + O2
hnu + ?
show the reaction diagram
-
assay at pH 7.6, about 100fold reduced relative luminescence
-
-
?
et-coelenterazine + O2
?
show the reaction diagram
f-coelenterazine + O2
hnu + ?
show the reaction diagram
-
assay at pH 7.6, about 100fold reduced relative luminescence
-
-
?
fcp-coelenterazine + O2
hnu + ?
show the reaction diagram
-
assay at pH 7.6, about 100fold reduced relative luminescence
-
-
?
h-coelenterazine + O2
?
show the reaction diagram
h-coelenterazine + O2
hnu + ?
show the reaction diagram
-
assay at pH 7.6m, about 100fold reduced relative luminescence
-
-
?
hcp-coelenterazine + O2
hnu + ?
show the reaction diagram
-
assay at pH 7.6, about 100fold reduced relative luminescence
-
-
?
i-coelenterazine + O2
?
show the reaction diagram
luciferin + O2
?
show the reaction diagram
-
assay at pH 7.4, 25-30°C, 10 min
-
-
?
me-coelenterazine + O2
?
show the reaction diagram
meo-coelenterazine + O2
?
show the reaction diagram
MeO-coelenterazine + O2
hnu + ?
show the reaction diagram
-
assay at pH 7.6, about 100fold reduced relative luminescence
-
-
?
n-coelenterazine + O2
hnu + ?
show the reaction diagram
-
assay at pH 7.6, about 100fold reduced relative luminescence
-
-
?
Renilla luciferin + O2
oxidized Renilla luciferin + CO2 + hv
show the reaction diagram
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
coelenterazine + O2
coelenteramide + CO2 + hv
show the reaction diagram
-
-
-
-
?
Renilla luciferin + O2
oxidized Renilla luciferin + CO2 + hv
show the reaction diagram
-
-
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Br-
-
stimulates luminescence activity
Cl-
-
stimulates luminescence activity
Cu2+
-
strong inhibition of luminescence activity
I-
-
stimulates luminescence activity
KCl
-
stimulation, optimum at 0.5 M
Mg2+
-
aequorin binds to Mg2+ under physiological conditions which modulates its light emission. The binding of Mg2+ to aequorin prevents the molecule from aggregating and stabilizes it in the monomeric form. Aequorin has two Mg2+ binding sites located within the EF-hands
NaCl
-
stimulation, optimum at 0.5 M
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(2Z)-3-[(2-bromophenyl)amino]-1-pyridin-2-ylprop-2-en-1-one
-
-
(2Z)-3-[[4-(dimethylamino)cyclohexa-1,5-dien-1-yl]amino]-1-phenylprop-2-en-1-one
-
-
(3Z)-3-[[4-(dimethylamino)phenyl]methylidene]-1,3-dihydro-2H-indol-2-one
-
common name SU4312
2-(4-ethoxyphenyl)-4-[(4-methylpiperazin-1-yl)carbonyl]quinoline
-
-
2-(4-ethoxyphenyl)-4-[(4-pyridin-2-ylpiperazin-1-yl)carbonyl]quinoline
-
-
2-(4-methylphenyl)-4-[(4-pyrimidin-2-ylpiperazin-1-yl)carbonyl]quinoline
-
-
2-(5-naphthalen-2-yl-1,2,4-oxadiazol-3-yl)pyridine
-
-
2-acetylaminofluorene
Renilla sp.
-
-
2-aminoanthracene
Renilla sp.
-
-
2-Benzylamidopyrazine
-
-
2-hydroxy-N'-[(1E)-(2-hydroxyphenyl)methylidene]benzohydrazide
-
common name SCS
2-mercaptoethanol
2-nitrofluorene
Renilla sp.
-
-
2-[5-(3-chlorophenyl)-1,2,4-oxadiazol-3-yl]pyridine
-
-
4-(1,4-dioxa-8-azaspiro[4.5]dec-8-ylcarbonyl)-2-(4-ethoxyphenyl)quinoline
-
-
4-acetylaminofluorene
Renilla sp.
-
-
4-amino-6-[(E)-[4'-[(Z)-(8-amino-1-hydroxy-5,7-disulfonato-6,7-dihydronaphthalen-2-yl)diazenyl]-3-methylbiphenyl-4-yl]diazenyl]-5-hydroxy-2,3-dihydronaphthalene-1,3-disulfonate
-
common name Evans Blue
4-nitroquinoline-N-oxide
Renilla sp.
-
-
4-[1-(1,3-benzothiazol-2-yl)-2-(4-methylpiperazin-1-yl)-2-oxoethyl]phenol
-
-
6-methyl-2-[(Z)-phenyldiazenyl]pyridin-3-ol
-
common name SIB1757
acetaminophen
Renilla sp.
-
-
actinomycin D
Renilla sp.
-
-
albendazole
Renilla sp.
-
-
amiodarone
Renilla sp.
-
-
ara C
Renilla sp.
-
-
arsenic III oxide
Renilla sp.
-
-
Benzyl luciferyl sulfate
-
-
Benzyl oxyluciferin
-
-
benzylamine
-
-
bleomycin
Renilla sp.
-
-
brefeldinA
-
5 microg/ml, 24 h, 90% decrease of Gluc secretion
carbamzepine
Renilla sp.
-
-
chlorambucil
Renilla sp.
-
-
chlorpromazine
Renilla sp.
-
-
cisplatin
Renilla sp.
-
-
coelenteramide
CuCl2
-
1 microM, inhibition of 95%
cyclophosphamide
Renilla sp.
-
-
cyclosporin A
Renilla sp.
-
-
cytochalasin B
Dibenzyl amine
-
-
diethyldicarbonate
-
-
digoxin
Renilla sp.
-
-
dimethyl sulfoxide
-
competitive inhibition at 16.6-66 mM, binding structure analysis by circular dichroism and fluorescence spectroscopy. Seven DMSO molecules interact with amino acids onthe surface of Renilla luciferase. Two of them interact with two catalytic residues (Glu144, His285), the rest of the DMSO molecules have specific interactions with the residues in the substrate binding site including Pro220, Phe180, and Phe261
dimethylnitrosamine
Renilla sp.
-
-
dithiothreitol
ethyl 4-[[2-(4-ethoxyphenyl)quinolin-4-yl]carbonyl]piperazine-1-carboxylate
-
-
ethyl methanesulfonate
Renilla sp.
-
-
Etioluciferin
-
-
etoposide
Renilla sp.
-
-
gemfibrozil
Renilla sp.
-
-
H-89
Renilla sp.
-
10 microM, reduced luminescence to 50% of RLuc and to 54% of RLuc8
haloperidol
Renilla sp.
-
-
Isopropanol
-
compatitive inhibition at 19.3-76 mM, binding structure analysis by circular dichroism and fluorescence spectroscopy. Four isopropanol molecules interact with amino acids. Most of these molecules move around the amino acidin the binding sites, and only one isopropanol molecule interacts with His285
Luciferyl sulfate
-
-
methyl oxyluciferin
-
-
methylmethane sulfonate
Renilla sp.
-
-
mitomycin
Renilla sp.
-
-
monensin A
-
3 microg/ml, 24 h, 65% decrease of Gluc secretion
N'-(3-chlorophenyl)-N-[(1Z)-(3-chlorophenyl)methylidene]imidoformamide
-
common name DCB
N-benzyl-4-methylbenzenesulfonamide
-
inhibition of purified enzyme, inducing activity in cell-based assay
N-methyl-N'-nitro-nitroso-guanidin
Renilla sp.
-
-
nefazodone
Renilla sp.
-
-
nocodazole
-
10 microg/ml, 24 h, 75% decrease of Gluc secretion
oxyluciferin
-
competitive
p-Anidisine
-
-
-
p-Benzyloxyaniline
-
-
phenytoin
Renilla sp.
-
-
Sodium azide
Renilla sp.
-
-
tacrine
Renilla sp.
-
-
taxol
Renilla sp.
-
-
thapsigargin
-
0.03 microM, about 30% decrease of Gluc secretion and activity
troglitazone
Renilla sp.
-
-
tyramine
-
-
Valproic acid
Renilla sp.
-
-
vinblastin
Renilla sp.
-
-
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(1,8-bis-maleimidotriethylene glycol)
Renilla sp.
-
effective conjugation of the N-terminal and C-terminal fragments of Rluc, results in enzymatic activity
-
ATP
Renilla sp.
-
50 microM
bovine serum albumin
-
addition to reaction mixture enhances luciferase activity
-
coelenterazine
-
increase of wild-type Gluc activity at higher substrate concentrations
histamine
Renilla sp.
-
10 microM
Triton X-100
-
0.1%, 3.7fold increase of specific activity of wild-tpye Gluc
additional information
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0016 - 0.0133
coelenterazine
0.0039
coelenterazine h
-
pH 7.8, 25°C, recombinant His-tagged enzyme
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.85
benzylluciferin
-
-
-
0.983
benzylluciferin methyl ether
-
-
-
3.9 - 4.9
coelenterazine
1.85
Luciferin
-
-
0.0167
methyl luciferin
-
-
-
additional information
coelenterazine
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.000022
coelenteramide
-
-
15.1
dimethyl sulfoxide
-
pH 7.8, 25°C, recombinant His-tagged enzyme
12.4
Isopropanol
-
pH 7.8, 25°C, recombinant His-tagged enzyme
0.0012
N-benzyl-4-methylbenzenesulfonamide
-
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.05
(2Z)-3-[(2-bromophenyl)amino]-1-pyridin-2-ylprop-2-en-1-one
Renilla reniformis;
-
IC50 above 0.05 mM
0.011
(2Z)-3-[[4-(dimethylamino)cyclohexa-1,5-dien-1-yl]amino]-1-phenylprop-2-en-1-one
Renilla reniformis;
-
; pH 7.2, 22°C
0.019
(3Z)-3-[[4-(dimethylamino)phenyl]methylidene]-1,3-dihydro-2H-indol-2-one
Renilla reniformis;
-
; pH 7.2, 22°C
0.025
2-(4-ethoxyphenyl)-4-[(4-methylpiperazin-1-yl)carbonyl]quinoline
Renilla reniformis;
-
IC50 above 0.025 mM
0.011
2-(4-ethoxyphenyl)-4-[(4-pyridin-2-ylpiperazin-1-yl)carbonyl]quinoline
Renilla reniformis;
-
; pH 7.2, 22°C
0.025
2-(4-methylphenyl)-4-[(4-pyrimidin-2-ylpiperazin-1-yl)carbonyl]quinoline
Renilla reniformis;
-
IC50 above 0.025 mM
0.025
2-(5-naphthalen-2-yl-1,2,4-oxadiazol-3-yl)pyridine
Renilla reniformis;
-
; pH 7.2, 22°C
0.02
2-hydroxy-N'-[(1E)-(2-hydroxyphenyl)methylidene]benzohydrazide
Renilla reniformis;
-
; pH 7.2, 22°C
0.05
2-[5-(3-chlorophenyl)-1,2,4-oxadiazol-3-yl]pyridine
Renilla reniformis;
-
IC50 above 0.05 mM
0.011
4-(1,4-dioxa-8-azaspiro[4.5]dec-8-ylcarbonyl)-2-(4-ethoxyphenyl)quinoline
Renilla reniformis;
-
; pH 7.2, 22°C
0.01
4-amino-6-[(E)-[4'-[(Z)-(8-amino-1-hydroxy-5,7-disulfonato-6,7-dihydronaphthalen-2-yl)diazenyl]-3-methylbiphenyl-4-yl]diazenyl]-5-hydroxy-2,3-dihydronaphthalene-1,3-disulfonate
Renilla reniformis;
-
-
0.0225
4-[1-(1,3-benzothiazol-2-yl)-2-(4-methylpiperazin-1-yl)-2-oxoethyl]phenol
Renilla reniformis;
-
; pH 7.2, 22°C
0.036
6-methyl-2-[(Z)-phenyldiazenyl]pyridin-3-ol
Renilla reniformis;
-
-
0.02
ethyl 4-[[2-(4-ethoxyphenyl)quinolin-4-yl]carbonyl]piperazine-1-carboxylate
Renilla reniformis;
-
; pH 7.2, 22°C
0.021
H-89
Renilla sp.;
-
10 microM coelenterazine, measured RLuc8 activity
0.05
N'-(3-chlorophenyl)-N-[(1Z)-(3-chlorophenyl)methylidene]imidoformamide
Renilla reniformis;
-
IC50 above 0.05 mM
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20
-
assay at
36 - 37
-
engineered enzyme mutant super RLuc8
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20 - 50
-
activity range, purified enzyme in absence of trehalose or sucrose
20 - 60
-
activity range, purified enzyme in presence of 1.2 M trehalose or sucrose
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
-
fraction in Escherichia coli cells
-
Manually annotated by BRENDA team
additional information
PDB
SCOP
CATH
UNIPROT
ORGANISM
D7PM14
Clytia gregaria;
Obelia longissima;
Renilla reniformis;
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
10350
-
MALDI-TOF-MS analyses, hGL-98/168
10740
-
MALDI-TOF-MS analyses, hGL-27/97
12000
-
2 * or 3 * 12000, SDS-PAGE
16000
Renilla sp.
-
value about, SDS-PAGE, C-terminal fragment
19000
-
catalytic domain
20020
-
MALDI-TOF-MS analyses, hGL
21610
-
calculated from sequence
25000
-
SDS-PAGE, pCold-hGL
30000
Renilla sp.
-
value about, SDS-PAGE, N-terminal fragment
33000 - 38000
36800
1 * 36800, mass spectrometry, 1 * 36900, calculated
36900
1 * 36800, mass spectrometry, 1 * 36900, calculated
37000
-
1 * 37000, SDS-PAGE, amino acid composition
37600
-
SDS-PAGE
37700
-
calculated from sequence
38040
-
calculated from sequence, 4red-shift luciferase
44000
-
SDS-PAGE
47000
-
Western blot, fusion protein Gluc-YFP
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer or trimer
-
2 * or 3 * 12000, SDS-PAGE
monomer
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
additional information
-
substitution of native Gaussia luciferase signal sequence by that from human interleukin-2 or albumin, decrease in luciferase activity
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
microbatch method is used for crystallization. The crystals of Ca2+-loaded apoaequorin are grown from 0.02 M CaCl2, 30% v/v 2-methyl-2,4-pentanediol, and 0.1 M sodium acetate (pH 4.6) during less than 1 week of incubation at 4°C. The maximum size of crystals is 0.35 * 0.3 * 0.25 mm
free enzyme and in complex with coelenterazine,and mutant K25A/E277A. Diffraction to 1.4 A. Structures demonstrate a classic alpha/beta-hydrolase fold. The presumptive catalytic triad residues are D120, E144, and H285. Additionally determination of structure of the accessory green fluorescent protein
molecular docking simulations with the coelenterazine substrate. Two triads of residues are critical for catalysis. Putative catalytic triad residues D120, E144, and H285 bear only limited resemblance to those of aequorin. Residues N53, W121, and P220 are also involved in catalysis
-
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 10
-
-
439334
6.5 - 8.5
-
below 6.5 and above 8.5 less than 20% residual activity
700221
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 50
-
at 50°C after 30 min incubation time only 20% residual activity, at 40°C after 1 h incubation time complete loss of activity
30
-
Rluc remaining activity in the absence and presence of 0.6 M and 1.2 M of trehalose, and 1.2 M of sucrose at 30°C is 15%, about 50%, around 100%, and almost 60%, respectively, after 60 min
30 - 52
-
the F116L/I137V mutant shows a transition starting at 30°C, which is 5°C lower than the wild-type, and ending at 52°C, which is 5°C higher than the wild type. N178D mutant has almost the same denaturation profile
35 - 47
-
purified recombinant His-tagged wild-type RLuc retains full activity up to 35°C and is inactivated at 47°C
39
Q6J4T7;
temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock, mutant enzyme F149S
40
-
in the absence and presence of 0.6 M of trehalose, the remaining activity is only about 2% and 6%, respectively, although it increases about 70fold upon incubation of the enzyme solution in a buffer containing 1.2 M of trehalose. It also seems that the remaining activity in the presence of 1.2 M of sucrose in the first 10 min is slightly better than in its absence, and a little activity is seen even after 60 min
40 - 47
-
the stability of the purified recombinant His-tagged N264S/S287P mutant is maintains at a temperature that is 5°C higher than the wild-type.
44
Q6J4T7;
temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock, wild-type enzyme and mutant enzyme E35G
46
Q6J4T7;
temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock, mutant enzyme E128G
48
Q6J4T7;
temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock, mutant enzyme L170I
49
Q6J4T7;
temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock, mutant enzyme D117G
50 - 70
-
engineered enzyme C-SRLuc8 shows significant stability, retaining about 20% of its initial activity after incubation at 70°C for 5 min, while wild-type SRLuc8 completely loses its activity after incubation at this temperature
51
Q6J4T7;
temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock, mutant enzyme D153G
52
Q6J4T7;
temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock, mutant enzyme Q168AR/L170M, Q168A/L170V and Q168K/L170V
53
Q6J4T7;
temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock, mutant enzyme Q168R
57
Q6J4T7;
temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock, mutant enzymes Q68R/L170I and Q168R/L179V
65
-
the residual activity of modified enzyme C-SRLuc8 is approximately 20% after incubation at 65°C for 5 min
95
-
3 min in the presence of DTT and then cooling on ice, over 93% of the luminescence activity of BFP-aq with coelenterazine is recovered
additional information
Renilla sp.
-
at 37°C reduced activity compared with remaining activity at 25°C
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
trehalose a thermostabilizing effect, a wide radial like network of trehalose molecules supports alpha-helix structures that are located in the N-terminus and C-terminus of the protein. In the water simulation box, these helices alter to instable structures at high temperatures. Reduction of the fluctuation of these helices in the presence of trehalose molecules, may prevent the protein from unfolding and increase its shelf-life
-
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
anaerobic luciferin binding promotes a rapid concentration-dependent aggregation of luciferase, which results in irreversible inactivation of the enzyme
-
439337
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4°C or -80°C, BFP-aq and gFP-aq do not show significant loss of their fluorescence and luciferase activities after 1 year
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
by affinity chromatography
-
Hi-Trap column
Renilla sp.
-
recombinant His-tagged engineered enzyme C-SRLuc8 in Escherichia coli by nickel affinity chromatography
-
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
-
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21 Star (DE3) by nickel affinity chromatography to over 95% purity
-
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
-
recombinant N-terminally His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and dialysis
-
recombinant soluble His6-tagged enzyme FcUni-RLuc from Escherichia coli strain BL21 (DE3) by nickel affinity chromatography
-
souble and insoluble fraction by nickel-chelate column
-
using Ni-chelate column
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
catalytic domain (19 kDa) is expressed using a cold-induced expression system in Escherichia coli as His-tagged fusion proteins
-
expressed in Escherichia coli
-
expressed in HEK-293T cells; expression of RLuc8 in Escherichia coli BL21
Renilla sp.
-
expressed using a cold-induced expression system in Escherichia coli as His-tagged fusion proteins
expression in 293T HEK fibroblasts and HF8 human fibroblasts
-
expression in Arabidopsis; expression in Escherichia coli strain BL21
-
expression in Chlamydomonas reinhardtii
expression in CHO cells and HepG2 cells
-
expression in CHO-K1 cells and PC12D cells
-
expression in Escherichia coli
expression in Escherichia coli BL21
-
expression in Escherichia coli DH10B cells and 293T human kidney fibroblasts
-
expression in Escherichia coli DH5alpha
-
expression in Escherichia coli strain DH5alpha and MCF-7 cells
Renilla sp.
-
expression in Escherichia coli strain ER 2566
Renilla sp.
-
expression in HEK-293 cells or Grip-Tite 293 MSR cells
-
expression in HeLa-cell
-
expression in Hep-G2 cell
Renilla sp.
-
expression in LNCaP cell, pRL-SV40 cell, pRL-null cell
-
expression in mouse and human C57/BL6 primary T cells, expression in PG-13 cells
expression in rabbit leukocyte lysate
Renilla sp.
-
expression of His-tagged apoclytin into the periplasmic space in Escherichia coli
-
expression of Renilla luciferase split fragments in HeLa cells, mitochondrial localization, overview
-
expression of stabilized variant RLuc8 with noncleavable N-terminal His6-tag
fragments of Renilla luciferase (Rluc) are fused to the chemotaxis-associated response regulator CheY3 and its phosphatase CheZ in the enteric pathogen Vibrio cholerae in order to demonstrate dynamic protein-protein interactions by luciferase complementation
-
gene luc, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3)
-
recombinant enzyme expression in ES-2 cells
-
recombinant expression of engineered His-tagged enzyme C-SRLuc8 in Escherichia coli, method optimization
-
recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3)
-
recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21 Star (DE3)
-
recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
-
recombinant expression of N-terminally His-tagged enzyme in Escherichia coli strain BL21(DE3)
-
recombinant expression of soluble His6-tagged enzyme FcUni-RLuc in Escherichia coli strain BL21 (DE3), an Fc-binding peptide is separated by a five-amino acid linker from RLuc
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D117G
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 5°C higher than wild-type value
D153G
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 7°C higher than wild-type value
E128G
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 2°C higher than wild-type value
E35G
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is identical to wild-type value
F149S
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 5°C lower than wild-type value
L170I
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 4°C higher than wild-type value
Q168A/L170V
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 8°C higher than wild-type value
Q168K/L170V
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 8°C higher than wild-type value
Q168R
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 9°C higher than wild-type value
Q168R/L170I
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 13°C higher than wild-type value
Q168R/L170M
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 8°C higher than wild-type value
Q168R/L170V
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 13°C higher than wild-type value
V44A
Q6J4T7;
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 1°C higher than wild-type value
DBHsp-GLuc
-
dopamine beta-hydroxlase fused to GLase, lower luminescence activity than wild-type GLuc
deltaSP-GLuc
-
GLuc without signal peptide sequence, no significant luminescence
extGLuc
-
membrane-anchored
Gluc(HPG)mutant
-
contains L-homopropargylglycine, prolonged luminescence, reduced specific activity
M43I
-
after 120 s of incubation time still 87% residual light emission compared to only 30% of wild-type Gluc, stabilized light emission with detergent Triton X-100, more stable light emission compared to wild-type Gluc when expression in mammalian cells
pCold-hGL
-
two repeat sequences, two catalytic domains
pCold-hGL-27/97
-
one repeat sequence, with only one catalytic domain
pCold-hGL-98/168
-
one repeat sequence, with only one catalytic domain
pCold-hSGL
-
with a signal peptide sequence for secretion
A55T/C124A/C130A/A143M/M253L/S287L/A123T/D154M/E155G/D162E/I163L/V185L
-
sequentially introduced into the RM-Luc coding sequence using designed oligonucleotide primers and quick-change site-directed mutagenesis, the mutant RM-Y has a red-shifted bioluminescence spectrum
A164W
-
73% of wild-type activity
A55T/C124A/S130A/K136R/A143M/M185V/M253L/S287L
-
selected mutations enable the protein to emit stronger bioluminescence activity and to be more stable in serum media. Mutant m-Rluc8 exhibits an enhancement in protein expression and shows a 5.6fold improvement in light output, with increased stability in serum media confirmed to last for over 5 days
D120E
-
1.1% of wild-type activity
D120F
-
no activity detected
D120Y
-
no activity detected
E144D
-
5.6% of wild-type activity
E144F
-
no activity detected
E144Y
-
no activity detected
E160N
-
27.2% of wild-type activity
F116L
-
random mutagenesis
F116L/I137V
-
random mutagenesis, solubility and specific activity of the mutant is higher compared to the wild-type
F180C
-
14.3% of wild-type activity
F180T
-
5.4% of wild-type activity
F180Y
-
11.0% of wild-type activity; 61.8% of wild-type activity
F261A
-
no activity detected
F261S
-
no activity detected
H285A
-
11.3% of wild-type activity
H285D
-
no activity detected
H285K
-
no activity detected
H285N
-
0.1% of wild-type activity
I137V
-
random mutagenesis
I140L
-
113% of wild-type activity
I163F
-
11.0% of wild-type activity
I223W
-
0.2% of wild-type activity
K189D
-
24.7% of wild-type activity
K189V
-
increased activity
K193S
-
54.8% of wild-type activity
K25A/E277A
surface mutations made with the intention that they would aid in crystallization, not involved in contacts between proteins in the crystal
K308I
-
47.5% of wild-type activity
M185V
-
increased activity
M185V/K189V/V267I
-
site-directed mutagenesis,compared with the native RLuc, mutant super RLuc has a higher turnover number, increased light output upon expression in Arabidopsis thaliana and enhanced half-life of photon emission, super RLuc is a blue light emitting luciferase
N178D
-
random mutagenesis, solubility and specific activity of the mutant is higher compared to the wild-type
N264S
-
random mutagenesis
N264S/S287P
-
random mutagenesis, solubility and specific activity of the mutant is higher compared to the wild-type
N45C/A71C
-
site-directed mutagenesis at the N-terminal of the enzyme, the engineered luciferase C-SRLuc8, improvement of the stability of super Renilla luciferase 8 (SRLuc8), which is a red-emitter variety of RLuc at higher temperatures, by introduction of a disulfide bridge into its structure. Evaluation of the proper disulfide bond formation based on computational methods, structure-function analysis, overview. The kinetic stability of C-SRLuc8 increases significantly at 60°C to 70°C as compared to SRLuc8. The N45C/A71C crosslink in C-SRLuc8 is involved in a hotspot foldon which seems to be the rate-limiting step of conformational collapse at higher temperatures. Molecular dynamic simulation studies to analyze the molecular basis of the structural changes after the introduction of the disulfide bridge. Increasing the local stability of several regions at this domain significantly improves the kinetic stability of C-SRLuc8, but the disulfide bridge in C-SRLuc8 does not delay the initial temperature of enzyme inactivation. The results of the thermal inactivation at 37°C and 65°C indicate that although CSRLuc8 shows a slight increase in stability during the first thirty minutes of incubation at 37°C, C-SRLuc8 shows a significant increase in thermostability at 65°C and increased activity as compared with SRLuc8
N53C
-
3.4% of wild-type activity
N53G
-
0.5% of wild-type activity
N53H
-
2.1% of wild-type activity
N53M
-
1.8% of wild-type activity
N53P
-
no activity detected
N53Q
-
25.1% of wild-type activity
N53R
-
90% of wild-type activity
N53S
-
20.7% of wild-type activity
P157R
-
101% of wild-type activity
P220C
-
72.7% of wild-type activity
P220E
-
4.9% of wild-type activity
P220F
-
15.7% of wild-type activity
P220M
-
140% of wild-type activity
P220Q
-
222% of wild-type activity
P220S
-
55.4% of wild-type activity
P220T
-
89.6% of wild-type activity
P220V
-
70.5% of wild-type activity
S287P
-
random mutagenesis
T184C
-
62.7% of wild-type activity
T184F
-
46.1% of wild-type activity
T329G
-
no significant influence on enzyme activity
V267I
-
increased activity
W121A
-
26.8% of wild-type activity
W121G
-
4.9% of wild-type activity
W121R
-
1.1% of wild-type activity
W121S
-
17.3% of wild-type activity
W121Y
-
3.1% of wild-type activity
223-224insRev
Renilla sp.
-
Rev peptide-inserted Rluc variant
223-224insTat
Renilla sp.
-
Tat peptide-inserted Rluc variant
223C/A-224insRev
Renilla sp.
-
Tat peptide-inserted Rluc variant
223C/A-224insTat
Renilla sp.
-
Tat peptide-inserted Rluc variant
229-230insRev
Renilla sp.
-
Rev peptide-inserted Rluc variant
229-230insTat
Renilla sp.
-
high thermal stability
229C/A-230insRev
Renilla sp.
-
Rev peptide-inserted Rluc variant
229C/A-230insTat
Renilla sp.
-
high thermal stability, higher enzyme activity than PI-RLuc
91-92C/AinsRev
Renilla sp.
-
higher enzyme activity than PI-RLuc
91-92insRev
Renilla sp.
-
Rev peptide-inserted Rluc variant
91-92insTat
Renilla sp.
-
higher enzyme activity than PI-RLuc
A55T/C124A/S130A/K136R/A143M/M185V/M253L/S287L
Renilla sp.
-
mutant enzyme exhibits a greater than 4-fold enhancement in activity, a 200fold increased resistance to serum inactivation, and a small but measurable 5 nm red shift in the emission spectrum. The enhancement in light output arises from a combination of increases in quantum yield and improvedkinetics
C124A
Renilla sp.
-
increased enzymatic activity, 41fold higher luminescence than wild-type luciferase
C124Ac
Renilla sp.
-
C-terminal half, decreased enzymatic activity
C124An
Renilla sp.
-
N-terminal half, decreased enzymatic activity
C126A
Renilla sp.
-
increased activity
extFLuc
Renilla sp.
-
membrane-anchored
G229stopstop
Renilla sp.
-
Rluc N-terminal fragment
M185V/Q235A
Renilla sp.
-
compared with the native enzyme the mutant has twice the rate of inactivation, as measured in murine serum, while incorporating a close to 5fold improvement in light output
N64C
Renilla sp.
-
Ca2+-induced interaction between calmodulin and M13 leads to intermolecular complementation of split Renilla luciferase, decreased enzymatic activity
RL8
Renilla sp.
-
mutant of Rluc
RLuc8
Renilla sp.
-
eight-point mutation variant of renilla luciferase
stop230
Renilla sp.
-
Rluc C-terminal fragment
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
biotechnology
diagnostics
molecular biology
pharmacology
additional information