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D117G
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 5°C higher than wild-type value
D153G
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 7°C higher than wild-type value
E128G
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 2°C higher than wild-type value
E35G
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is identical to wild-type value
F149S
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 5°C lower than wild-type value
L170I
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 4°C higher than wild-type value
Q168A/L170V
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 8°C higher than wild-type value
Q168K/L170V
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 8°C higher than wild-type value
Q168R
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 9°C higher than wild-type value
Q168R/L170I
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 13°C higher than wild-type value
Q168R/L170M
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 8°C higher than wild-type value
Q168R/L170V
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 13°C higher than wild-type value
V44A
the temperature for 50% inactivation of aequorin bioluminescence by a 30-min heat shock is 1°C higher than wild-type value
DBHsp-GLuc
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dopamine beta-hydroxlase fused to GLase, lower luminescence activity than wild-type GLuc
deltaSP-GLuc
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GLuc without signal peptide sequence, no significant luminescence
extGLuc
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membrane-anchored
Gluc(HPG)mutant
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contains L-homopropargylglycine, prolonged luminescence, reduced specific activity
M43I
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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
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two repeat sequences, two catalytic domains
pCold-hGL-27/97
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one repeat sequence, with only one catalytic domain
pCold-hGL-98/168
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one repeat sequence, with only one catalytic domain
pCold-hSGL
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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
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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
F116/I137V
the mutant starts to denature at 30°C, and retained its activity up to 52°C with increased solubility at 34°C and specific activity up to approximately 119%
F116L/I137V
random mutagenesis, solubility and specific activity of the mutant is higher compared to the wild-type
F116L/I137V/I75A/N178D/N264S/S287P
the thermostability effect increases, with the mutant showing approximately 10°C higher stability. The mutant shows improved tolerance for protease digestion, e.g. trypsin and proteinase and for organic solvent. The mutant enzyme retains 100% specific activity at 45°C, while the wild type loses almost all activity, and retains activity at 55°C. The specific activity is approximately 123% higher than that of the wild type
F116L/I137V/N264S/S287P
thermostability of the mutant is increased. The mutant enzyme shows denaturation at 45 to 52°C and specific activity up to approximately 150% compared with the wild type enzyme
F180C
14.3% of wild-type activity
F180T
5.4% 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
I140L
113% of wild-type activity
I163F
11.0% of wild-type activity
I223W
0.2% of wild-type activity
I75A
specific activity of I75A is 47% of that of the wild type enzyme, retains activity up to 50°C
K189D
24.7% of wild-type activity
K189V
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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
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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
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
T184C
62.7% of wild-type activity
T184F
46.1% of wild-type activity
T329G
-
no significant influence on enzyme activity
V267I
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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.
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Rev peptide-inserted Rluc variant
223-224insTat
Renilla sp.
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Tat peptide-inserted Rluc variant
223C/A-224insRev
Renilla sp.
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Tat peptide-inserted Rluc variant
223C/A-224insTat
Renilla sp.
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Tat peptide-inserted Rluc variant
229-230insRev
Renilla sp.
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Rev peptide-inserted Rluc variant
229-230insTat
Renilla sp.
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high thermal stability
229C/A-230insRev
Renilla sp.
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Rev peptide-inserted Rluc variant
229C/A-230insTat
Renilla sp.
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high thermal stability, higher enzyme activity than PI-RLuc
91-92C/AinsRev
Renilla sp.
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higher enzyme activity than PI-RLuc
91-92insRev
Renilla sp.
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Rev peptide-inserted Rluc variant
91-92insTat
Renilla sp.
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higher enzyme activity than PI-RLuc
A55T/C124A/S130A/K136R/A143M/M185V/M253L/S287L
Renilla sp.
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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.
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increased enzymatic activity, 41fold higher luminescence than wild-type luciferase
C124Ac
Renilla sp.
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C-terminal half, decreased enzymatic activity
C124An
Renilla sp.
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N-terminal half, decreased enzymatic activity
C126A
Renilla sp.
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increased activity
extFLuc
Renilla sp.
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membrane-anchored
G229stopstop
Renilla sp.
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Rluc N-terminal fragment
M185V/Q235A
Renilla sp.
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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.
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Ca2+-induced interaction between calmodulin and M13 leads to intermolecular complementation of split Renilla luciferase, decreased enzymatic activity
RL8
Renilla sp.
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mutant of Rluc
RLuc8
Renilla sp.
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eight-point mutation variant of renilla luciferase
stop230
Renilla sp.
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Rluc C-terminal fragment
F180Y
11.0% of wild-type activity
F180Y
61.8% of wild-type activity
M185G
16.7% of wild-type activity
M185G
-
slightly increased half life
N178D
random mutagenesis, solubility and specific activity of the mutant is higher compared to the wild-type
N178D
mutation does not affect thermostability but increases the solubility at 34°C and specific activity up to approximately 141%
N264S/S287P
random mutagenesis, solubility and specific activity of the mutant is higher compared to the wild-type
N264S/S287P
the mutant starts to denature at 40°C and retains its activity up to 50°C with increased solubility at 34°C and specific activity up to approximately 150%
P220G
548% of wild-type activity
P220G
-
only 4% of the initial luciferase activity of wild-type luciferase
P220L
500% of wild-type activity
P220L
-
only 16% of the initial luciferase activity of wild-type luciferase
additional information
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conjugate of GLuc with the IM9 protein
additional information
construct of vector with signal peptide from GLuc and gene sequence from human endostatin
additional information
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construct of vector with signal peptide from GLuc and gene sequence from human endostatin
additional information
fusion of GLuc to a biotin acceptor peptide, attachment of a single biotin to the fusion protein mediated by the biotin protein ligase, EC 6.3.4.15
additional information
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fusion of GLuc to a biotin acceptor peptide, attachment of a single biotin to the fusion protein mediated by the biotin protein ligase, EC 6.3.4.15
additional information
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fusion of Gluc to fluorescent protein YFP
additional information
substitution of native Gaussia luciferase signal sequence by that from human interleukin-2 or albumin, reduced amount of protein, decreased luciferase activity
additional information
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substitution of native Gaussia luciferase signal sequence by that from human interleukin-2 or albumin, reduced amount of protein, decreased luciferase activity
additional information
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without the signal sequence higher enzyme yield and activity
additional information
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functional enzyme secreted by mammalian cells due to fusion to the signal peptide of human interleukin-2
additional information
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mutation Cys152Ala in fusion to signal peptide of human interleukin-2 stabilizes
additional information
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analysis of the Hsp90 chaperone activity in complex with cochaperone Cdc37 using split Renilla luciferase protein fragment-assisted complementation bioluminescence, full-length human Hsp90-Cdc37 complex and critical residues contributing to Hsp90/Cdc37 interaction in living cells, computational modeling and molecular dynamics simulations, overview. Cysteines at the N-terminus of Cdc37 do not directly contribute to Hsp90-Cdc37 complex formation
additional information
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method development of Renilla luciferase used in an assay for measurement of mitochondrial fusion, quantification via split-Renilla luciferase complementation in HeLa cells, overview
additional information
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substitution of the V3 region of the multifunctional NS5A protein of hepatitis C virus FH1 with the Renilla reniformis luciferase Rluc gene. The deletion of the V3 region from the genome does only slightly affect the titer of infectious virus produced in human hepatoma cell line, Huh 7.5. The transfected virus stably expresses an NS5A-Rluc fusion protein, kinetics of virus production, overview
additional information
enzyme engineering of blue light emitting enzyme mutant super RLuc to construct a luciferase with desired light emission wavelength and thermostability, namely super RLuc8, which has a red-shifted spectrum and shows stable light emission. Super RLuc8 shows a 10fold increase in thermostability at 37°C after 20 min incubation, in comparison to the native enzyme. The optimum temperature of the mutant increases from 30 to 37°C. Molecular dynamics simulation analysis indicates that the increased thermostability is most probably caused by a better structural compactness and more local rigidity in the regions out of the emitter site. The mutant super RLuc8 shows increased activity compared t the wild-type. Molecular dynamics simulation, overview
additional information
overall structure of the MU-RLuc model involving five mutated residues, F116L, I137V, N178D, N264S and S287P, overview
additional information
stabilization of luciferase from Renilla reniformis using random mutations
additional information
super RLuc8 is a Renilla luciferase variant in which 16 amino acids are substituted
additional information
the enzyme is fused to wild-type LC3 protein and LC3 mutant G120A. LC3-II turnover is used as a marker of autophagic flux. The Rluc-LC3 fusion protein is used for the indirect autophagy flux assay based on monitoring the degradation of an autophagosome-associated fusion protein Rluc-LC3 by luminescence detection. The Rluc-LC3 assay is useful for the identification of genes, miRNAs, and small molecules that regulate autophagy flux in mammalian cells. Design of the RLUC-LC3wt and RLUC-LC3G120A fusion proteins, recombinantly expressed in MCF-7 cells. Method evaluation and optimization, overview