Ansamycin Antibiotics as Free Radical Scavengers Isolated from Streptomyces by Using the Bactericidal Action of the Hydroxyl Radical

Ansamycin antibiotics (1–4) were isolated from the cultured broth of Streptomyces sp. USF-319 strain as a result of our screening for free radical scavengers. They inhibited the bactericidal effect of the Fenton reagent toward Bacillus subtilis by their radical scavenging activity. Some of them also showed inhibitory activity against lipid peroxidation and lipoxygenases.     

A new method for rapid identification of ansamycin compounds by inactivating KLM gene clusters in potential ansamycin-producing actinomyces

Aims: In this study, we explored the possibility of construction of a ‘universal targeting vector’ by Red/ET recombination to inactivate L gene encoding 3‐amino‐5‐hydroxybenzoic acid (AHBA)‐oxidoreductase in AHBA biosynthetic gene cluster to facilitate the detection of ansamycins production in actinomycetes. Methods and Results: Based on the conserved regions of linked AHBA synthase (K), oxidoreductase (L) and phosphatase (M) gene clusters, degenerate primers were designed and PCR was performed to detect KLM gene clusters within 33 AHBA synthase gene‐positive actinomycetes strains. Among them, 22 KLM gene cluster‐positive strains were identified. A ‘universal targeting vector’ was further constructed using the 50‐nt homologous sequences chosen from four strains internal L gene in KLM gene clusters through Red/ET recombination. The L gene from nine of the KLM gene cluster‐positive actinomycetes strains was inactivated by insertion of a kanamycin (Km) resistance marker into its internal region from the ‘universal targeting vector’. By comparison of the metabolites produced in parent strains with those in L gene‐inactivated mutants, we demonstrated the possible ansamycins production produced by these strains. One strain (4089) was proved to be a geldanamycin producer. Three strains (3‐20, 7‐32 and 8‐32) were identified as potential triene‐ansamycins producers. Another strain (3‐27) was possible to be a streptovaricin C producer. Strains 24‐100 and 4‐124 might be served as ansamitocin‐like producers. Conclusions: The results confirmed the feasibility that a ‘universal targeting vector’ could be constructed through Red/ET recombination using the conserved regions of KLM gene clusters to detect ansamycins production in actinomycetes. Significance and Impact of the Study: The ‘universal targeting vector’ provides a rapid approach in certain degree to detect the potential ansamycin producers from the 22 KLM gene cluster‐positive actinomycetes strains.     

PCR screening reveals unexpected antibiotic biosynthetic potential in Amycolatopsis sp. strain UM16

AIMS: To assess the antibiotic biosynthetic potential of Amycolatopsis sp. strain UM16 and eight other Amycolatopsis species. METHODS AND RESULTS: Amycolatopsis genomic DNA was screened by PCR for the glycopeptide, Type-II (aromatic) polyketide and ansamycin biosynthetic gene clusters. Amycolatopsis sp. strain UM16, which exhibits weak antitubercular activity, was shown to have the glycopeptide oxyB gene and the Type-II (aromatic) polyketide-synthase KSalpha-KSbeta tandem gene pair, but not the AHBA synthase gene. The ristocetin (glycopeptide) producer, Amycolatopsis lurida NRRL 2430(T), was shown to have the oxyB gene and the Type-II polyketide-synthase KSalpha-KSbeta tandem gene pair. Amycolatopsis alba NRRL 18532(T) was shown to have the glycopeptide oxyB gene and the AHBA synthase gene. Phylogenetic analyses using Amycolatopsis oxyB and KSalpha-KSbeta gene sequences were conducted. CONCLUSIONS: Amycolatopsis sp. strain UM16 appears to have the biosynthetic potential to produce glycopeptide and Type-II polyketide antibiotics, but not ansamycins. The potential to synthesize aromatic polyketides may be more widely distributed in Amycolatopsis than is currently recognized. SIGNIFICANCE AND IMPACT OF THE STUDY: PCR screening is a very useful tool for rapidly identifying the biosynthetic potential of an antibiotic-producing actinomycete isolate. Advanced knowledge of the type of antibiotic(s) produced will allow appropriate methods to be selected for antibiotic purification.     

Hsp90: a specialized but essential protein-folding tool

Hsp90 is unique among molecular chaperones. The majority of its known substrates are signal transduction proteins, and recent work indicates that it uses a novel protein-folding strategy.     

Protoplast preparation and regeneration in Nocardia mediterranei

Abstract A standard procedure for the preparation of viable Streptomyces protoplasts was found to be unsuccessful for the rifamycin-producing actinomycete Nocardia mediterranei . An alternative method for N. mediterranei protoplast preparation is described. This method was also found to be applicable to several other ansamycin-producing actinomycetes.     

New Olimycins from a Cold-Seep-Derived Streptomyces olivaceus

Cold-seeps are areas of the ocean floor in which hydrogen sulfide and methane are released into the open water. The cold-seep microbes are an emerging source of novel bioactive natural products. Four new ansa-ring opened linear ansamycin analogues, named olimycins E−H ( 1 – 4 ) were isolated from the cold-seep-derived Streptomyces olivaceus OUCLQ19-3. The planar and stereochemical structures of the isolated compounds were elucidated based on extensive MS and NMR spectroscopic analyses together with ECD calculations.