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Information on EC 1.13.12.5 - Renilla-type luciferase

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IUBMB Comments
This enzyme has been studied from the soft coral Renilla reniformis. Before the reaction occurs the substrate is sequestered by a coelenterazine-binding protein. Elevation in the concentration of calcium ions releases the substrate, which then interacts with the luciferase. Upon binding the substrate, the enzyme catalyses an oxygenation, producing a very short-lived hydroperoxide that cyclizes into a dioxetanone structure, which collapses, releasing a CO2 molecule. The spontaneous breakdown of the dioxetanone releases the energy (about 50 kcal/mole) that is necessary to generate the excited state of the coelenteramide product, which is the singlet form of the monoanion. In vivo the product undergoes the process of nonradiative energy transfer to an accessory protein, a green fluorescent protein (GFP), which results in green bioluminescence. In vitro, in the absence of GFP, the product emits blue light.
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UNIPROT: Q9BLZ2
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Word Map
The enzyme appears in viruses and cellular organisms
Reaction Schemes
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Synonyms
aequorin, renilla luciferase, gaussia luciferase, gaussia princeps luciferase, renilla reniformis luciferase, rluc8, clytin, oplophorus luciferase, bfp-aq, gaussia-luciferase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Caussia princeps luciferase
-
Gaussia luciferase
-
Gaussia-luciferase
-
aequorin
-
-
-
-
luciferase (Renilla luciferin)
-
-
-
-
Renilla luciferin 2-monooxygenase
-
-
-
-
Renilla-luciferin 2-monooxygenase
-
-
-
-
Renilla-type luciferase
-
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oxidative decarboxylation
-
redox reaction
-
-
-
-
oxidation
-
-
-
-
reduction
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
coelenterazine h:oxygen 2-oxidoreductase (decarboxylating)
This enzyme has been studied from the soft coral Renilla reniformis. Before the reaction occurs the substrate is sequestered by a coelenterazine-binding protein. Elevation in the concentration of calcium ions releases the substrate, which then interacts with the luciferase. Upon binding the substrate, the enzyme catalyses an oxygenation, producing a very short-lived hydroperoxide that cyclizes into a dioxetanone structure, which collapses, releasing a CO2 molecule. The spontaneous breakdown of the dioxetanone releases the energy (about 50 kcal/mole) that is necessary to generate the excited state of the coelenteramide product, which is the singlet form of the monoanion. In vivo the product undergoes the process of nonradiative energy transfer to an accessory protein, a green fluorescent protein (GFP), which results in green bioluminescence. In vitro, in the absence of GFP, the product emits blue light.
CAS REGISTRY NUMBER
COMMENTARY hide
346421-46-3
Gaussia luciferase
61869-41-8
-
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
coelenterazine + O2
?
show the reaction diagram
0.1 mM
-
-
?
coelenterazine + O2
? + CO2 + hv
show the reaction diagram
-
-
-
?
coelenterazine + O2
coelenteramide + CO2 + hnu
show the reaction diagram
-
-
-
?
coelentrazine + O2
?
show the reaction diagram
-
-
-
?
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
coelenteramide
-
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
KCl
substitution of NaCl by KCl does not significantly alter the dependence of light intensity on salt concentrations
NaCl
maximum activity at 1.0-1.5 M, a high NaCl concentration enhances luciferase stability to thermal denaturation
additional information
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00214
coelenterazine
K0.5 value, Hill coefficient 1.8, pH 7.5, temperature not specified in the publication
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
9
60% residual activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
1
30% residual activity
37
20% residual activity
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
enzyme shows positive cooperativity with coelenterazine, rather due to the kinetic cooperativity conditioned by conformational changes in response to substrate binding than to the presence of two catalytic sites
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION?
Q9BLZ2_9MAXI
185
0
19900
TrEMBL
-
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
18800
SDS-PAGE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
x * 19062, MALDI-TOF, x * 19056, calcuated from sequence
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
side-chain modification
additional information
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
solution structure of fully active, recombinant GLuc. GLuc is an all-alphahelix protein made of nine helices. Two homologous sequential repeats form two anti-parallel bundles made by 4 helices and tied together by three disulfide bonds. The N-terminal helix 1 is grabbed by these 4 helices
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
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
72
melting temperature
77
melting temperature, presence of 1 M NaCl
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20°C, 50% glycerol, stable for months
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
by affinity chromatography
natively folded GLuc is obtained by bacterial expression and efficient refolding using a solubility enhancement petide tag
recombinant secreted enzyme
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Chlamydomonas reinhardtii
expression in CHO cells and HepG2 cells
expression in insect cells using a baculovirus expression system
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Verhaegen, M.; Christopoulos, T.
Recombinant Gaussia luciferase: Overexpression, purification, and analytical application of a bioluminescent reporter for DNA hybridization
Anal. Chem.
74
4378-4385
2002
Gaussia princeps (Q9BLZ2), Gaussia princeps
Manually annotated by BRENDA team
Knappskog, S.; Ravneberg, H.; Gjerdrum, C.; Tre, C.; Stern, B.; Pryme, I.
The level of synthesis and secretion of Gaussia princeps luciferase in transfected CHO cells is heavily dependent on the choice of signal peptide
J. Biotechnol.
128
705-715
2007
Gaussia princeps (Q9BLZ2), Gaussia princeps
Manually annotated by BRENDA team
Ruecker, O.; Zillner, K.; Groebner-Ferreira, R.; Heitzer, M.
Gaussia-luciferase as a sensitive reporter gene for monitoring promoter activity in the nucleus of the green alga Chlamydomonas reinhardtii
Mol. Genet. Genomics
280
153-162
2008
Renilla sp., Gaussia princeps (Q9BLZ2), Gaussia princeps
Manually annotated by BRENDA team
Broyles, D.B.; Dikici, E.; Daunert, S.; Deo, S.K.
Facile synthesis and characterization of a novel tamavidin-luciferase reporter fusion protein for universal signaling applications
Adv. Biosyst.
4
e1900166
2020
Gaussia princeps (Q9BLZ2)
Manually annotated by BRENDA team
Larionova, M.; Markova, S.; Vysotski, E.
Bioluminescent and structural features of native folded Gaussia luciferase
J. Photochem. Photobiol. B
183
309-317
2018
Gaussia princeps (Q9BLZ2), Gaussia princeps
Manually annotated by BRENDA team
Wu, N.; Kobayashi, N.; Tsuda, K.; Unzai, S.; Saotome, T.; Kuroda, Y.; Yamazaki, T.
Solution structure of Gaussia Luciferase with five disulfide bonds and identification of a putative coelenterazine binding cavity by heteronuclear NMR
Sci. Rep.
10
20069
2020
Gaussia princeps (Q9BLZ2)
Manually annotated by BRENDA team
Blower, I.; Tong, C.; Sun, X.; Murray, E.; Luckett, J.; Chan, W.; Williams, P.; Hill, P.
Gaussia luciferase as a reporter for quorum sensing in Staphylococcus aureus
Sensors
20
4305
2020
Gaussia princeps (Q9BLZ2)
Manually annotated by BRENDA team