Biopolym. Cell. 2002; 18(2):91-95.
Construction and selecthion strains Streptomyces peucetius subsp. caesius ATCC 27952-2 with elevated DXR production
1Dubitska L. P., 2Fedorenko V. O.
  1. Danylo Halytsky Lviv National Medical University
    69, Pekarska Str., Lviv, Ukraine, 79010
  2. Ivan Franko National University of L'viv
    4, Hrushevskoho Str., Lviv, Ukraine, 79005


Genes cpx and dnrl (encode P-450 enzyme that hydroxilize daunorubicin (DNR) converting it to doxorubicin (DXR) and transcription activator, respectively) leukaemomycin biosynthesis gene cluster of S. griseus were cloned in multicopy pIJ486 under control of strong promoter not (from noursethricin resistance gen noursei) and introduced into strains S. peucetius subsp. caesius ACC 27952-2 a TK24. 48-hous old cultures of strains 279S2-2 (pIJ486cpx dnrl+), TK24 (pIJ486cpx+ dnrl DNRr-mutants of strain 27952-2 synthesize DXR from exogenously added DNR. In s 27952-2 pJJ486cpx+ dnrl production of antracydines, DNR and DXR was increased. Although guantity of converted DNR and concentration of P-450 hydroxylase was highest in DNRr-mutants of strain 27952-2. Thus, resistance of DNRr-mutants to DNR depends not only on functioning of res gene but also on conversion of more toxic for given strain compound (DNR) to I (DXR). It allows to obtain more valuable antibiotic from its Intermediate.


[1] Bulkina ZP. Anticancer drugs. Kiev: Naukova Dumka, 1991. 304 p.
[2] Cabriales S, Bresnahan J, Testa D, Espina BM, Scadden DT, Ross M, Gill PS. Extravasation of liposomal daunorubicin in patients with AIDS-associated Kaposi's sarcoma: a report of four cases. Oncol Nurs Forum. 1998;25(1):67-70.
[3] Cassinelli G, Arlandini E, Ballabio M, Bordoni T, Geroni C, Giuliani F, Grein A, Merli S, Rivola G. New biosynthetic anthracyclines related to barminomycins incorporating barbiturates in their moiety. J Antibiot (Tokyo). 1990;43(1):19-28.
[4] Hutchinson CR. Biosynthetic Studies of Daunorubicin and Tetracenomycin C. Chem Rev. 1997;97(7):2525-2536.
[5] Crameri R, Davies JE. Increased production of aminoglycosides associated with amplified antibiotic resistance genes. J Antibiot (Tokyo). 1986;39(1):128-35.
[6] Retzlaff L, Mayer G, Beyer S, Ahlert J, Verseck S, Distler J, Piepersbery W. Industrial microorganisms: Basic and applied molecular genetics. Eds R. H. Baltz, G. D. Hegeman, P. L. Skatrud. Washington: Amer. Soc. Microbiol. Publ., 1993: 183-94.
[7] Dubytska LP, Fedorenko VO. Characteristic of mutants of Streptomyces peucetius subsp. caesius ATCC 27952 resistant to anthracycline antibiotics. Mikrobiol Z. 2001; 63(4):53-61.
[8] Pat. USA 5955319. Process for preparing doxorubicin. L. Fonstein., S. Otten., C. Hutchinson. Publ. 1999.
[9] Pat. USA 5364781. Process for preparing daunorubicin. C. R. Hutchinson., K. M. Madduri., F. Torti., A. L. Colombo. Publ. 1994.
[10] Pat. USA 592293. Methods of producing doxorubicin. W. R. Strohl., M. L. Dickens., C. R. Desanti. Publ. 1999.
[11] Dickens ML, Strohl WR. 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. 1996;178(11):3389-95.
[12] Gauze GF, Preobrazhenskaya TP, Sveshnikova MA, Terekhova MP, Maksimova TS. Actinomycetes. M.: Nauka, 1983. 244 p.
[13] Hopwood DA, Bibb NJ, Chater KF, Kieser T, Bruton CJ, Kieser HM, Lidiate D, Smith CP, Ward JM, Schrempf H. Genetic manipulation of Streptomyces: a laboratory manual. Norwich: The John Innes Fund., 1985. 365 p.
[14] Segura D, Santana C, Gosh R, Escalante L, Sanchez S. Anthracyclines: isolation of overproducing strains by the selection and genetic recombination of putative regulatory mutants of Streptomycespeucetius var. caesius. Appl Microbiol Biotechnol. 1997;48(5):615–20.
[15] Dornberger K, Stengel C, Miosga N. Thin-layer chromatographic analysis of adriamycinone in fermentation broths. J Chromatogr A. 1985;328:432–5.
[16] Omura T, Sato R. The carbon monoxide-binding pigment of liver microsomes. I. Evidence for its hemoprotein nature. J Biol Chem. 1964;239:2370-8.
[17] Lowry OH, Rosebrough NJ, Farr Al, Randall RJ. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951;193(1):265-75.
[18] Maniatis T, Fritsch EF, Sambrook J. Molecular cloning: a laboratory manual. New York: Cold Spring Harbor Lab, 1982; 545 p.
[19] Kaur P. Expression and characterization of DrrA and DrrB proteins of Streptomyces peucetius in Escherichia coli: DrrA is an ATP binding protein. J Bacteriol. 1997;179(3):569-75.
[20] Walczak RJ, Dickens ML, Priestley ND, Strohl WR. Purification, properties, and characterization of recombinant Streptomyces sp. strain C5 DoxA, a cytochrome P-450 catalyzing multiple steps in doxorubicin biosynthesis. J Bacteriol. 1999;181(1):298-304.
[21] Stutzman-Engwall KJ, Otten SL, Hutchinson CR. Regulation of secondary metabolism in Streptomyces spp. and overproduction of daunorubicin in Streptomyces peucetius. J Bacteriol. 1992;174(1):144-54.
[22] Seno ET, Cox KL. Maintenance of cloned tylosin biosynthetic genes in S. fradiae on freely-replicating and integrative plasmid vectors. J Cell Biochem. 1990. 14A: 93.