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Protein

Cytochrome P-450 monooxygenase DoxA

Gene

doxA

Organism
Streptomyces sp. (strain C5)
Status
Reviewed-Annotation score: Annotation score: 5 out of 5-Experimental evidence at protein leveli

Functioni

Involved in the biosynthesis of the anthracyclines carminomycin and daunorubicin (daunomycin) which are aromatic polyketide antibiotics that exhibit high cytotoxicity and are widely applied in the chemotherapy of a variety of cancers. In vivo, DoxA catalyzes the C-13 hydroxylation of 13-deoxycarminomycin and 13-deoxydaunorubicin to yield 13-dihydrocarminomycin and 13-dihydrodaunorubicin, respectively, as well as the oxidation of these 13-dihydro-anthracyclines to their respective 13-keto forms, carminomycin and daunorubicin. In vitro, it also catalyzes the C-14 hydroxylation of daunorubicin to form doxorubicin (adriamycin), although this strain is not a doxorubicin producer. It is not able to accept anthracyclinones (aglycones) and anthracyclines with a 10-carbomethoxyl moiety. 13-oxidation of the anthracyclines possessing the 4-methoxy substitution is greatly favored. The anthracycline analog desacetyladriamycin can be oxidized to 10-hydroxydesacetyladriamycin. It can only use NADP. DoxA acts jointly with DauV.4 Publications

Catalytic activityi

13-deoxydaunorubicin + NADPH + O2 = 13-dihydrodaunorubicin + NADP+ + H2O.4 Publications
13-dihydrodaunorubicin + NADPH + O2 = daunorubicin + NADP+ + 2 H2O.4 Publications
13-deoxycarminomycin + NADPH + O2 = 13-dihydrocarminomycin + NADP+ + H2O.4 Publications
13-dihydrocarminomycin + NADPH + O2 = carminomycin + NADP+ + H2O.4 Publications

Cofactori

hemeBy similarity

Enzyme regulationi

Strongly inhibited by dithiothreitol and high ionic strength buffers.1 Publication

Kineticsi

Kcat is 0.63 sec(-1) for oxidation on 13-dihydrodaunorubicin. Kcat is 0.024 sec(-1) for hydroxylation on 13-deoxydaunorubicin. Kcat is 0.00026 sec(-1) for oxidation on 13-dihydrocarminomycin. Kcat is 0.00012 sec(-1) for hydroxylation on daunorubicin.

  1. KM=0.9 µM for daunorubicin (at pH 7.5 and 30 degrees Celsius)1 Publication
  2. KM=1.1 µM for 13-deoxydaunorubicin (at pH 7.5 and 30 degrees Celsius)1 Publication
  3. KM=2.5 µM for 13-dihydrocarminomycin (at pH 7.5 and 30 degrees Celsius)1 Publication
  4. KM=4.6 µM for 13-dihydrodaunorubicin (at pH 7.5 and 30 degrees Celsius)1 Publication
  1. Vmax=7.5 pmol/min/mg enzyme with daunorubicin as substrate (at pH 7.5 and 30 degrees Celsius)1 Publication
  2. Vmax=19 pmol/min/mg enzyme with 13-dihydrocarminomycin as substrate (at pH 7.5 and 30 degrees Celsius)1 Publication
  3. Vmax=1600 pmol/min/mg enzyme with 13-deoxydaunorubicin as substrate (at pH 7.5 and 30 degrees Celsius)1 Publication
  4. Vmax=3900 pmol/min/mg enzyme with 13-dihydrodaunorubicin as substrate (at pH 7.5 and 30 degrees Celsius)1 Publication

pH dependencei

Optimum pH is 7.5. This optimal pH, slightly higher than normal physiological conditions, may be necessary to ensure that the proper ionic nature of the zwitterionic anthracycline substrates is maintained for optimal recognition.1 Publication

Temperature dependencei

Optimum temperature is 30 degrees Celsius.1 Publication

Pathwayi: daunorubicin biosynthesis

This protein is involved in the pathway daunorubicin biosynthesis, which is part of Antibiotic biosynthesis.
View all proteins of this organism that are known to be involved in the pathway daunorubicin biosynthesis and in Antibiotic biosynthesis.

Pathwayi: carminomycin biosynthesis

This protein is involved in the pathway carminomycin biosynthesis, which is part of Antibiotic biosynthesis.
View all proteins of this organism that are known to be involved in the pathway carminomycin biosynthesis and in Antibiotic biosynthesis.

Sites

Feature keyPosition(s)DescriptionActionsGraphical viewLength
Metal bindingi369Iron (heme axial ligand)By similarity1

GO - Molecular functioni

GO - Biological processi

  • daunorubicin biosynthetic process Source: UniProtKB
Complete GO annotation...

Keywords - Molecular functioni

Monooxygenase, Oxidoreductase

Keywords - Biological processi

Antibiotic biosynthesis

Keywords - Ligandi

Heme, Iron, Metal-binding, NADP

Enzyme and pathway databases

BioCyciMetaCyc:MONOMER-18172.
BRENDAi1.14.13.181. 1284.
UniPathwayiUPA00054.
UPA01040.

Names & Taxonomyi

Protein namesi
Recommended name:
Cytochrome P-450 monooxygenase DoxA (EC:1.14.13.1814 Publications)
Alternative name(s):
13-deoxycarminomycin C-13 hydroxylase
13-deoxydaunorubicin C-13 hydroxylase
13-dihydrocarminomycin C-13 hydroxylase
13-dihydrodaunorubicin C-13 hydroxylase
Gene namesi
Name:doxA
OrganismiStreptomyces sp. (strain C5)
Taxonomic identifieri45212 [NCBI]
Taxonomic lineageiBacteriaActinobacteriaStreptomycetalesStreptomycetaceaeStreptomyces

Subcellular locationi

GO - Cellular componenti

Complete GO annotation...

Keywords - Cellular componenti

Cytoplasm

PTM / Processingi

Molecule processing

Feature keyPosition(s)DescriptionActionsGraphical viewLength
ChainiPRO_00004256831 – 422Cytochrome P-450 monooxygenase DoxAAdd BLAST422

Interactioni

Subunit structurei

Monomer.1 Publication

Structurei

3D structure databases

ProteinModelPortaliQ59971.
SMRiQ59971.
ModBaseiSearch...
MobiDBiSearch...

Family & Domainsi

Sequence similaritiesi

Belongs to the cytochrome P450 family.Curated

Phylogenomic databases

KOiK15955.

Family and domain databases

Gene3Di1.10.630.10. 1 hit.
InterProiIPR001128. Cyt_P450.
IPR002397. Cyt_P450_B.
IPR017972. Cyt_P450_CS.
[Graphical view]
PfamiPF00067. p450. 1 hit.
[Graphical view]
PRINTSiPR00359. BP450.
PR00385. P450.
SUPFAMiSSF48264. SSF48264. 1 hit.
PROSITEiPS00086. CYTOCHROME_P450. 1 hit.
[Graphical view]

Sequencei

Sequence statusi: Complete.

Q59971-1 [UniParc]FASTAAdd to basket

« Hide

        10         20         30         40         50
MSGEAPRVAV DPFSCPMMTM QRKPEVHDAF REAGPVVEVN APAGGPAWVI
60 70 80 90 100
TDDALAREVL ADPRFVKDPD LAPTAWRGVD DGLDIPVPEL RPFTLIAVDG
110 120 130 140 150
EDHRRLRRIH APAFNPRRLA ERTDRIAAIA DRLLTELADS SDRSGEPAEL
160 170 180 190 200
IGGFAYHFPL LVICELLGVP VTDPAMAREA VGVLKALGLG GPQSAGGDGT
210 220 230 240 250
DPAGDVPDTS ALESLLLEAV HAARRKDTRT MTRVLYERAQ AEFGSVSDDQ
260 270 280 290 300
LVYMITGLIF AGHDTTGSFL GFLLAEVLAG RLAADADGDA ISRFVEEALR
310 320 330 340 350
HHPPVPYTLW RFAATEVVIR GVRLPRGAPV LVDIEGTNTD GRHHDAPHAF
360 370 380 390 400
HPDRPSRRRL TFGDGPHYCI GEQLAQLESR TMIGVLRSRF PQARLAVPYE
410 420
ELRWCRKGAQ TARLTDLPVW LR
Length:422
Mass (Da):46,096
Last modified:November 1, 1996 - v1
Checksum:iF4EADECA1D159052
GO

Sequence databases

Select the link destinations:
EMBLi
GenBanki
DDBJi
Links Updated
U50973 Genomic DNA. Translation: AAB08049.1.

Genome annotation databases

KEGGiag:AAB08049.

Cross-referencesi

Sequence databases

Select the link destinations:
EMBLi
GenBanki
DDBJi
Links Updated
U50973 Genomic DNA. Translation: AAB08049.1.

3D structure databases

ProteinModelPortaliQ59971.
SMRiQ59971.
ModBaseiSearch...
MobiDBiSearch...

Protocols and materials databases

Structural Biology KnowledgebaseSearch...

Genome annotation databases

KEGGiag:AAB08049.

Phylogenomic databases

KOiK15955.

Enzyme and pathway databases

UniPathwayiUPA00054.
UPA01040.
BioCyciMetaCyc:MONOMER-18172.
BRENDAi1.14.13.181. 1284.

Family and domain databases

Gene3Di1.10.630.10. 1 hit.
InterProiIPR001128. Cyt_P450.
IPR002397. Cyt_P450_B.
IPR017972. Cyt_P450_CS.
[Graphical view]
PfamiPF00067. p450. 1 hit.
[Graphical view]
PRINTSiPR00359. BP450.
PR00385. P450.
SUPFAMiSSF48264. SSF48264. 1 hit.
PROSITEiPS00086. CYTOCHROME_P450. 1 hit.
[Graphical view]
ProtoNetiSearch...

Entry informationi

Entry nameiDOXA_STRS5
AccessioniPrimary (citable) accession number: Q59971
Entry historyi
Integrated into UniProtKB/Swiss-Prot: March 19, 2014
Last sequence update: November 1, 1996
Last modified: November 2, 2016
This is version 73 of the entry and version 1 of the sequence. [Complete history]
Entry statusiReviewed (UniProtKB/Swiss-Prot)
Annotation programProkaryotic Protein Annotation Program

Miscellaneousi

Miscellaneous

Streptomyces sp. C5 produces significant quantities of doxorubicin only when doxA is strongly overexpressed, but under normal conditions, 14-hydroxylation of daunorubicin cannot favorably compete with baumycin biosynthesis from daunorubicin. S.peucetius subsp. caesius ATCC 27952, a mutant strain derived from S.peucetius ATCC 29050, is the only organism reported to produce doxorubicin in vivo (PubMed:9864343).1 Publication

Documents

  1. PATHWAY comments
    Index of metabolic and biosynthesis pathways
  2. SIMILARITY comments
    Index of protein domains and families

Similar proteinsi

Links to similar proteins from the UniProt Reference Clusters (UniRef) at 100%, 90% and 50% sequence identity:
100%UniRef100 combines identical sequences and sub-fragments with 11 or more residues from any organism into one UniRef entry.
90%UniRef90 is built by clustering UniRef100 sequences that have at least 90% sequence identity to, and 80% overlap with, the longest sequence (a.k.a seed sequence).
50%UniRef50 is built by clustering UniRef90 seed sequences that have at least 50% sequence identity to, and 80% overlap with, the longest sequence in the cluster.