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Information on EC 1.14.13.181 - 13-deoxydaunorubicin hydroxylase and Organism(s) Streptomyces sp. and UniProt Accession Q59971

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IUBMB Comments
The enzymes from the Gram-positive bacteria Streptomyces sp. C5 and Streptomyces peucetius show broad substrate specificity for structures based on an anthracycline aglycone, but have a strong preference for 4-methoxy anthracycline intermediates (13-deoxydaunorubicin and 13-dihydrodaunorubicin) over their 4-hydroxy analogues (13-deoxycarminomycin and 13-dihydrocarminomycin), as well as a preference for substrates hydroxylated at the C-13 rather than the C-14 position.
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Streptomyces sp.
UNIPROT: Q59971
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The taxonomic range for the selected organisms is: Streptomyces sp.
The enzyme appears in selected viruses and cellular organisms
Synonyms
cyp109a2, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
DoxA monooxygenase
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SYSTEMATIC NAME
IUBMB Comments
13-deoxydaunorubicin,NADPH:oxygen oxidoreductase (13-hydroxylating)
The enzymes from the Gram-positive bacteria Streptomyces sp. C5 and Streptomyces peucetius show broad substrate specificity for structures based on an anthracycline aglycone, but have a strong preference for 4-methoxy anthracycline intermediates (13-deoxydaunorubicin and 13-dihydrodaunorubicin) over their 4-hydroxy analogues (13-deoxycarminomycin and 13-dihydrocarminomycin), as well as a preference for substrates hydroxylated at the C-13 rather than the C-14 position.
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
13-deoxycarminomycin + NADPH + H+ + O2
13-dihydrocarminomycin + NADP+ + H2O
show the reaction diagram
13-deoxydaunorubicin + NADPH + H+ + O2
13-dihydrodaunorubicin + NADP+ + H2O
show the reaction diagram
13-deoxydaunorubicin + NADPH + H+ + O2
daunorubicinol + NADP+ + H2O
show the reaction diagram
-
-
-
?
13-dihydrocarminomycin + NADPH + H+ + O2
carminomycin + NADP+ + H2O
show the reaction diagram
13-dihydrodaunorubicin + NADPH + H+ + O2
daunorubicin + NADP+ + 2 H2O
show the reaction diagram
13-dihydrodaunorubicin + NADPH + H+ + O2
daunorubicin + NADP+ + H2O
show the reaction diagram
DoxA catalyzes three steps in the daunorubicin/doxorubicin biosynthesis pathway: hydroxylation at C13 of 13-deoxycarminomycin and 13-deoxydaunorubicin, oxidation at C-13 of 13-dihydrocarminomycin and daunorubicinol and the hydroxylation at C-14 of daunorubicin. It appears that the primary function of DoxA is to catalyze the enzymatic conversion of 13-deoxydaunorubicin to daunorubicin via daunorubicinol
-
-
?
daunomycin + NADPH + H+ + O2
doxorubicin + NADP+ + H2O
show the reaction diagram
daunorubicin + NADPH + H+ + O2
doxorubicin + NADP+ + H2O
show the reaction diagram
additional information
?
-
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
13-deoxycarminomycin + NADPH + H+ + O2
13-dihydrocarminomycin + NADP+ + H2O
show the reaction diagram
13-deoxydaunorubicin + NADPH + H+ + O2
13-dihydrodaunorubicin + NADP+ + H2O
show the reaction diagram
13-dihydrocarminomycin + NADPH + H+ + O2
carminomycin + NADP+ + H2O
show the reaction diagram
13-dihydrodaunorubicin + NADPH + H+ + O2
daunorubicin + NADP+ + 2 H2O
show the reaction diagram
DoxA catalyzes three steps in the daunorubicin/doxorubicin biosynthesis pathway: hydroxylation at C13 of 13-deoxycarminomycin and 13-deoxydaunorubicin, oxidation at C-13 of 13-dihydrocarminomycin and daunorubicinol and the hydroxylation at C-14 of daunorubicin
-
-
?
13-dihydrodaunorubicin + NADPH + H+ + O2
daunorubicin + NADP+ + H2O
show the reaction diagram
DoxA catalyzes three steps in the daunorubicin/doxorubicin biosynthesis pathway: hydroxylation at C13 of 13-deoxycarminomycin and 13-deoxydaunorubicin, oxidation at C-13 of 13-dihydrocarminomycin and daunorubicinol and the hydroxylation at C-14 of daunorubicin. It appears that the primary function of DoxA is to catalyze the enzymatic conversion of 13-deoxydaunorubicin to daunorubicin via daunorubicinol
-
-
?
daunomycin + NADPH + H+ + O2
doxorubicin + NADP+ + H2O
show the reaction diagram
DoxA catalyzes three steps in the daunorubicin/doxorubicin biosynthesis pathway: hydroxylation at C13 of 13-deoxycarminomycin and 13-deoxydaunorubicin, oxidation at C-13 of 13-dihydrocarminomycin and 13-dihydrodaunorubicin and the hydroxylation at C-14 of daunorubicin
-
-
?
daunorubicin + NADPH + H+ + O2
doxorubicin + NADP+ + H2O
show the reaction diagram
DoxA catalyzes three steps in the daunorubicin/doxorubicin biosynthesis pathway: hydroxylation at C13 of 13-deoxycarminomycin and 13-deoxydaunorubicin, oxidation at C-13 of 13-dihydrocarminomycin and daunorubicinol and the hydroxylation at C-14 of daunorubicin. It appears that the primary function of DoxA is to catalyze the enzymatic conversion of 13-deoxydaunorubicin to daunorubicin via daunorubicinol
-
-
?
additional information
?
-
the enzyme convert daunomycin to doxorubicin
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
cytochrome P-450
cytochrome P-450-dependent monooxygenase
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NADPH
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
dithiothreitol
strongly inhibitory, inhibition is reversible after desalting
doxorubicin
completely inhibits DoxA activity at both 1 and 5 mM, competitive with respect to daunorubicinol oxidation
quinidine
inhibits DoxA oxidation of daunorubicinol by 11% at 1 mM. At 5 mM, it inhibits at more than 95%
sulfaphenazole
inhibits DoxA oxidation of daunorubicinol by 20% at 1 mM. At 5 mM, it inhibits at more than 95%
troleandomycin
inhibits DoxA oxidation of daunorubicinol by 35% at 1 mM. At 5 mM, it inhibits at more than 95%
additional information
high ionic strength buffers containing 100 mM sodium phosphate, also strongly inhibits DoxA activity. This effect is partially reversible after exchange into low ionic strength buffers, such as 20 mM N-(2-hydroxyethyl)piperazine-N9-(2-ethanesulfonic acid) (HEPES) or 10 mM sodium phosphate (pH 7.5). No inhibition: 4-methylpyrazole (1 or 5 mM), rhodomycin D (1-5 mM)
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0025
13-deoxycarminomycin
pH 7.5, 30°C
0.0011
13-deoxydaunorubicin
pH 7.5, 30°C
0.0046
13-dihydrodaunorubicin
pH 7.5, 30°C
0.0009
daunorubicin
pH 7.5, 30°C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00026
13-deoxycarminomycin
pH 7.5, 30°C
0.024
13-deoxydaunorubicin
pH 7.5, 30°C
0.63
13-dihydrodaunorubicin
pH 7.5, 30°C
0.00012
daunorubicin
pH 7.5, 30°C
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.104
13-deoxycarminomycin
pH 7.5, 30°C
21.81
13-deoxydaunorubicin
pH 7.5, 30°C
136.9
13-dihydrodaunorubicin
pH 7.5, 30°C
0.133
daunorubicin
pH 7.5, 30°C
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.021
doxorubicin
pH 7.5, 30°C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0006
pH 7.5, 30°C, 13-oxidation of daunorubicinol to daunorubicin
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
DoxA catalyzes three steps in the daunorubicin/doxorubicin biosynthesis pathway: hydroxylation at C13 of 13-deoxycarminomycin and 13-deoxydaunorubicin, oxidation at C-13 of 13-dihydrocarminomycin and 13-dihydrodaunorubicin and the hydroxylation at C-14 of daunorubicin. It appears that the primary function of DoxA is to catalyze the enzymatic conversion of 13-deoxydaunorubicin to daunorubicin via daunorubicinol
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
46096
47000
1 * 47000, SDS-PAGE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
x * 46096, calculated from sequence
monomer
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
22
stable for 1 h in presence of 20% (vol/vol) glycerol. All activity is lost after 30 min at room temperature in the absence of glycerol
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4°C, 20% (v/v) glycerol, 7 days, no appreciable loss of activity
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
the recombinant DoxA is purified to homogeneity from Streptomyces lividans transformed with a plasmid containing the Streptomyces sp. strain C5 doxA gene under the control of the strong SnpR-activated snpA promoter
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Streptomyces lividans TK24
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Dickens, M.L.; Strohl, W.R.
Isolation and characterization of a gene from Streptomyces sp. strain C5 that confers the ability to convert daunomycin to doxorubicin on Streptomyces lividans TK24
J. Bacteriol.
178
3389-3395
1996
Streptomyces sp. (Q59971)
Manually annotated by BRENDA team
Dickens, M.L.; Priestley, N.D.; Strohl, W.R.
In vivo and in vitro bioconversion of epsilon-rhodomycinone glycoside to doxorubicin: functions of DauP, DauK, and DoxA
J. Bacteriol.
179
2641-2650
1997
Streptomyces sp. (Q59971)
Manually annotated by BRENDA team
Walczak, R.J.; Dickens, M.L.; Priestley, N.D.; Strohl, W.R.
Purification, properties, and characterization of recombinant Streptomyces sp. strain C5 DoxA, a cytochrome P-450 catalyzing multiple steps in doxorubicin biosynthesis
J. Bacteriol.
181
298-304
1999
Streptomyces sp. (Q59971)
Manually annotated by BRENDA team