Subcloning and expression of midecamycin polyketide synthase genes from Streptomyces mycarofaciens 1748.
A 2.4 kb fragment containing the midecamycin polyketide synthase genes (mps) was subcloned from the preliminary clone pCN8B12 out of the genomic library of midecamycin-producing strain S. mycarofaciens 1748, by using the homologous DNA of the actinorhodin polyketide synthase gene (act I) as hybridization probe. This DNA fragment was subcloned onto Streptomyces/E. coli shuttle vector pMHM3. A recombinant plasmid pCG2 was obtained. The transformation of the polyketide synthase deficient mutant of actinorhodin-producing strain, S. colicolor TK17, with pCG2 DNA resulted in the production of an antibacterial compound which was similar neither to actinorhodin nor to midecamycin. The transformation of spiramycin-producing strain S. ambofaciens with pCG2 DNA increased spiramycin production in the fermentation broth. The transformation of the regulatory mutant of daunorubicin-producing strain with pCG2 DNA resulted in the production of epsilon-rhodomycinone verified by TLC and HPLC analyses. The pCG2 DNA also could be functionally expressed in tetracenomycin C-producing strain S. glaucescens. However, it could not be expressed in the blocked mutants of erythromycin-producing strain Saccharopolyspora erythrarea WMH 15,261. These suggest that the pCG2 DNA may complement polyketide synthase gene deficiency or have some regulatory function in certain polyketide antibiotic-producing strains.
Zhu
X
X
Institute of Medicinal Biotechnology, Chinese Academy of Medical Science, Beijing.
Wang
Y
Y
Jin
L
L
Xu
X
X
D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000032:analysis; D005798:Genes, Bacterial / Q000235:genetics*; D005822:Genetic Vectors; D007933:Leucomycins / Q000096:biosynthesis*; D009097:Multienzyme Complexes / Q000096:biosynthesis / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D013302:Streptomyces / Q000235:genetics*
D004269:DNA, Bacterial; D007933:Leucomycins; D009097:Multienzyme Complexes; D011994:Recombinant Proteins; C026483:midecamycin