title: [Ribosome engineering of streptomyces sp. FJ3 from Three Gorges reservoir area and metabolic product of the selected mutant strain].
abstract: OBJECTIVE
  To explore new resource from inactive actinomycete strains, we screened resistant mutant strains by ribosome engineering, and analyzed the products derived from the selected mutant strains.
  
  
  METHODS
  Three Gorges reservoir area-derived actinomycete strains including BD20, FJ3, WZ20 and FJ5 were used as initial strains, which showed no-antibacterial activities. The streptomycin-resistant (str(R)) mutants and rifampicin-resistant (rif(R)) mutants were screened by single colony isolation on streptomycin-containing plates and rifampicin-containing plates according to the method for obtaining drug-resistant mutants in ribosome engineering. The four initial strains and their str(R)-mutants and rif(R)-mutants were fermented in a liquid medium with the same composition. Mutants with anti-Staphylococcus aureus activity were obtained by paper chromatography. The components of fermentation broth were analyzed by high performance liquid chromatography (HPLC) and high performance liquid chromatography-mass spectrometry (LC-MS). Furthermore, FJ3 strain was identified by 16S rDNA and morphology.
  
  
  RESULTS
  The minimal inhibitory concentration (MIC) of streptomycin and rifampicin for FJ3 was: 0.5 microg/mL and 110 microg/mL, respectively. Twenty-four strR-mutant strains and 20 rif(R)-mutant strains of FJ3 mutant strains were selected for bioassay. The result of the antibacterial activity screening demonstrated that six strains inhibited bacteria. Two strains (FJ3-2 and FJ3-6) were screened from the streptomycin-resistance mutants of inactive strain FJ3. The result of bioassay showed that the fermentation broth of FJ3-2 and FJ3-6 exhibited obvious anti-Staphylococcus aureus activity. The assay of paper chromatography showed that the active substance may be nucleic acid class antibiotic via using solvent system Doskochilova. Moreover, the results of HPLC and LC-MS exhibited that this substance may be thiolutin.
  
  
  CONCLUSIONS
  Ribosome engineering for changing the secondary metabolic function of the inactive wild-type actinomycete strains was a feasible method for the acquirement of active mutant strains, which will be beneficial to exploit the new medical actinomycete strains.
authors: Southwest University, College of Pharmacy, Chongqing 400716, China. haile6@163.com
authors: Le
authors: L
authors: Hai
authors: Yuqi
authors: Y
authors: Huang
authors: Guojian
authors: G
authors: Liao
authors: Changhua
authors: C
authors: Hu
mesh_terms: D002851:Chromatography, High Pressure Liquid; D024881:Drug Resistance, Bacterial; D005818:Genetic Engineering; D013058:Mass Spectrometry; D008826:Microbial Sensitivity Tests; D009154:Mutation; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012270:Ribosomes / Q000378:metabolism*; D012293:Rifampin / Q000494:pharmacology; D013211:Staphylococcus aureus / Q000187:drug effects; D013302:Streptomyces / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D013307:Streptomycin / Q000494:pharmacology
chemical_list: D012336:RNA, Ribosomal, 16S; D012293:Rifampin; D013307:Streptomycin