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Characterization of aklavinone-11-hydroxylase from Streptomyces purpurascens.
- Source :
-
Biochimica et biophysica acta [Biochim Biophys Acta] 1999 Feb 10; Vol. 1430 (1), pp. 57-64. - Publication Year :
- 1999
-
Abstract
- Aklavinone-11-hydroxylase (RdmE) is a FAD monooxygenase participating in the biosynthesis of daunorubicin, doxorubicin and rhodomycins. The rdmE gene encodes an enzyme of 535 amino acids. The sequence of the Streptomyces purpurascens enzyme is similar to other Streptomyces aromatic polyketide hydroxylases. We overexpressed the gene in Streptomyces lividans and purified aklavinone-11-hydroxylase to apparent homogeneity with four chromatographic steps utilizing a kinetic photometric enzyme assay. The enzyme is active as the monomer with a molecular mass of 60 kDa; it hydroxylates aklavinone and other anthracyclinones. Aklavinone-11-hydroxylase can use both NADH and NADPH as coenzyme but it is slowly inactivated in the presence of NADH. The apparent Km for NADPH is 2 mM and for aklavinone 10 microM. The enzyme is inactivated in the presence of phenylglyoxal and 2,3-butanedione. NADPH protects against inactivation of aklavinone-11-hydroxylase by phenylglyoxal.
- Subjects :
- Anthracyclines pharmacokinetics
Aryl Hydrocarbon Hydroxylases antagonists & inhibitors
Aryl Hydrocarbon Hydroxylases chemistry
Biotransformation
Genes, Bacterial
Kinetics
NADP
Phenylglyoxal
Streptomyces genetics
Temperature
Aryl Hydrocarbon Hydroxylases isolation & purification
Streptomyces enzymology
Subjects
Details
- Language :
- English
- ISSN :
- 0006-3002
- Volume :
- 1430
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Biochimica et biophysica acta
- Publication Type :
- Academic Journal
- Accession number :
- 10082933
- Full Text :
- https://doi.org/10.1016/s0167-4838(98)00265-9