Streptomyces peucetius daunorubicin biosynthesis gene, dnrF: sequence and heterologous expression

• Published in: Microbiology (Society for General Microbiology) 1995-04, Vol.141 (4), p.1007-1016
• Main Authors: Filippini, Silvia, Solinas, Maria Manuela, Breme, Umberto, Schluter, Manuela Birgit, Gabellini, Davide, Biamonti, Giuseppe, Colombo, Anna Luisa, Garofano, Luisa
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Aryl Hydrocarbon Hydroxylases - genetics; Aryl Hydrocarbon Hydroxylases - metabolism; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bacteriology; Base Sequence; Biological and medical sciences; Biology of microorganisms of confirmed or potential industrial interest; Biotechnology; Chromosome Mapping; Cloning, Molecular; Daunorubicin - biosynthesis; DNA Primers - genetics; DNA, Bacterial - genetics; Escherichia coli; Escherichia coli - genetics; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Bacterial; Genes, Bacterial; Genetics; Microbiology; Mission oriented research; Molecular Sequence Data; Open Reading Frames; Sequence Homology, Amino Acid; Streptomyces - genetics; Streptomyces - metabolism; Streptomyces lividans; Streptomyces peucetius; Substrate Specificity
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/13500872-141-4-1007

BioScience Center PHARMACIA, Via Papa Giovanni XXIII 23, 20014 Nerviano (Mi), Italy Istituto di Genetica Biochimica ed Evoluzionistica del CNR, Via Abbiategrasso 207, 27100 Pavia, Italy ABSTRACT The dnrF gene, responsible for conversion of aklavinone to -rhodomycinone via C-11 hydroxylation, was mapped in the daunorubicin (Dnr) gene cluster of Streptomyces peucetius ATCC 29050, close to drrAB, one of the anthracycline-resistance genes. The dnrF gene was sequenced and should encode a protein of 489 amino acids with a molecular mass of 52 kDa. The deduced DnrF protein shows significant similarities with bacterial FAD- and NADPH-dependent hydroxylases either required to introduce hydroxyl groups into polycyclic aromatic polyketide antibiotics or involved in catabolism of aromatic compounds. Heterologous expression of dnrF in Streptomyces lividans TK23 and in Escherichia coli demonstrated that the gene encodes a NADPH-dependent hydroxylase catalysing the hydroxylation of aklavinone to yield -rhodomycinone. The enzyme is inactive on anthracyclines glycosylated at position C-7 and its activity decreases to a different extent with other substrate modifications, indicating that DnrF has a significant substrate specificity. Author for correspondence: Silvia Filippini. Tel: [phone]. Fax: [phone]. The GenBank accession number of the sequence reported in this paper is U18082