1.13.12.13: Oplophorus-luciferin 2-monooxygenase

This is an abbreviated version!
For detailed information about Oplophorus-luciferin 2-monooxygenase, go to the full flat file.

Word Map on EC 1.13.12.13

Reaction

Oplophorus luciferin
+
O2
=
oxidized Oplophorus luciferin
+
CO2
+
hnu

Synonyms

19kOLase, imidazopyrazinone-type luciferase, KAZ, luciferase, nanoKAZ, NanoLuc, Oluc-19, Oplophorus luciferase

ECTree

     1 Oxidoreductases
         1.13 Acting on single donors with incorporation of molecular oxygen (oxygenases)
             1.13.12 With incorporation of one atom of oxygen (internal monooxygenases or internal mixed-function oxidases)
                1.13.12.13 Oplophorus-luciferin 2-monooxygenase

Engineering

Engineering on EC 1.13.12.13 - Oplophorus-luciferin 2-monooxygenase

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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
L72Q
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
Y138I
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
V44I/Y138I
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows wild-type bioluminesce intensity
V44I/A54I/Y138I
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, catalytic subunit mutant, that shows 7fold higher activity than 16-aa-mutant nanoKAZ using coelenterazine, but these substitutions does not stimulate protein secretion from mammalian cells, nanoKAZ possessing the signal peptide sequence of Gaussia luciferase for secretion expressed efficiently into the culture medium of CHO-K1 cells
V44I/A54I
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows the mutant shows increased bioluminesce intensity compared to the wild-type
V44I
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
Q18L
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows wild-type bioluminesce intensity
Q124K
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
Q11R
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
P115E
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
N166R
M75K
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
A33N
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
L27V
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows reduced bioluminesce intensity compared to the wild-type
K43R
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows wild-type bioluminesce intensity
I90V
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
F68D
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows wild-type bioluminesce intensity
C164S
-
mutation does not significantly affect catalytic activity of subunit kOLase
C164G
-
mutation does not significantly affect catalytic activity of subunit kOLase
C164A
-
mutation does not significantly affect catalytic activity of subunit kOLase
A54I/Y138I
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
A54I
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows increased bioluminesce intensity compared to the wild-type
A4E/Q11R/Q18L/L27V/A33N/K43R/V44I/A54I/F68D/L72Q/M75K/I90V/P115E/Q124K/Y138I/N166R
Q9GV45; Q9GV46
construction of mutant nanoKAZ, a mutant with 16 amino acid substitutions of the catalytic subunit, by site-directed mutagenesis, the mutant nanoKAZ lacks the amino-terminal signal peptide, crystal structure determination and analysis, structure-function relationship in nanoKAZ, overview. The truncation of 10 amino acid residues at N- and C-terminal regions of nanoKAZ causes a complete loss of luminescence activity. Both the alpha1-helix and beta11-strand in the nanoKAZ molecule might serve to stabilize the molecule, which is essential in catalyzing the luminescence reaction
A4E/Q11R/A33K/V44I/A54F/P115E/Q124K/Y138I/Q18L/F54I/F68Y/L72Q/M75K/I90V/L27V/K33N/K43R/Y68D
-
mutant Nluc: Nluc paired with furimazine produces 2.5 millionfold brighter luminescence in mammalian cells relative to Oluc-19 with coelenterazine. The luminescence produced by Nluc decays with a half-life more than 2 h, significantly longer than for mutant A4E/Q11R/A33K/V44I/A54F/P115E/Q124K/Y138I. Nluc increased luminescence is gained mostly through improvements in protein stability, where Nluc shows markedly greater retention of activity in lysates following incubation at 37°C
A4E/Q11R/A33K/V44I/A54F/P115E/Q124K/Y138I
-
mutant is 29000fold brighter than mutant N166R. Western blot analysis shows that mutant A4E/Q11R/A33K/V44I/A54F/P115E/Q124K/Y138I is produced more efficiently than mutant N166R in cells. The increased expression is consistent with improved enzyme stability at 37°C, where the half-life of activity retention is increased 65fold over that of mutant N166R
A4E
Q9GV45; Q9GV46
site-directed mutagenesis, mutation of the catalytic subunit, the mutant shows wild-type bioluminesce intensity
additional information