Regio- and stereoselective intermolecular oxidative phenol coupling in Streptomyces.

Intermolecular oxidative phenol coupling is the main process in nature for the formation of atroposelective biaryl compounds. Although well defined in plants and fungi, this type of dimerization reaction in bacteria is poorly understood. Therefore, the biosynthesis of julichromes, spectomycins, and setomimycin was investigated. The monomeric subunits of these biarylic pre-anthraquinones are derived from a common polyketidic precursor, yet the coupling reaction proceeds in a regioselective manner, with the position of attachment of the two subunits depending on the specific streptomycete strain. By using genome analysis and deletion experiments, the biosynthetic gene clusters were identified. Furthermore, it was established that cytochrome P450 enzymes are fundamentally involved during dimerization of the polyketide monomers.

Tracking down biotransformation to the genetic level: identification of a highly flexible glycosyltransferase from Saccharothrix espanaensis.

Saccharothrix espanaensis is a member of the order Actinomycetales. The genome of the strain has been sequenced recently, revealing 106 glycosyltransferase genes. In this paper, we report the detection of a glycosyltransferase from Saccharothrix espanaensis which is able to rhamnosylate different phenolic compounds targeting different positions of the molecules. The gene encoding the flexible glycosyltransferase is not located close to a natural product biosynthetic gene cluster. Therefore, the native function of this enzyme might be not the biosynthesis of a secondary metabolite but the glycosylation of internal and external natural products as part of a defense mechanism.

Complete genome sequence of Saccharothrix espanaensis DSM 44229(T) and comparison to the other completely sequenced Pseudonocardiaceae.

BACKGROUND: The genus Saccharothrix is a representative of the family Pseudonocardiaceae, known to include producer strains of a wide variety of potent antibiotics. Saccharothrix espanaensis produces both saccharomicins A and B of the promising new class of heptadecaglycoside antibiotics, active against both bacteria and yeast. RESULTS: To better assess its capabilities, the complete genome sequence of S. espanaensis was established. With a size of 9,360,653 bp, coding for 8,501 genes, it stands alongside other Pseudonocardiaceae with large genomes. Besides a predicted core genome of 810 genes shared in the family, S. espanaensis has a large number of accessory genes: 2,967 singletons when compared to the family, of which 1,292 have no clear orthologs in the RefSeq database. The genome analysis revealed the presence of 26 biosynthetic gene clusters potentially encoding secondary metabolites. Among them, the cluster coding for the saccharomicins could be identified. CONCLUSION: S. espanaensis is the first completely sequenced species of the genus Saccharothrix. The genome discloses the cluster responsible for the biosynthesis of the saccharomicins, the largest oligosaccharide antibiotic currently identified. Moreover, the genome revealed 25 additional putative secondary metabolite gene clusters further suggesting the strain's potential for natural product synthesis.

Site-specific recombination strategies for engineering actinomycete genomes.

The feasibility of using technologies based on site-specific recombination in actinomycetes was shown several years ago. Despite their huge potential, these technologies mostly have been used for simple marker removal from a chromosome. In this paper, we present different site-specific recombination strategies for genome engineering in several actinomycetes belonging to the genera Streptomyces, Micromonospora, and Saccharothrix. Two different systems based on Cre/loxP and Dre/rox have been utilized for numerous applications. The activity of the Cre recombinase on the heterospecific loxLE and loxRE sites was similar to its activity on wild-type loxP sites. Moreover, an apramycin resistance marker flanked by the loxLERE sites was eliminated from the Streptomyces coelicolor M145 genome at a surprisingly high frequency (80%) compared to other bacteria. A synthetic gene encoding the Dre recombinase was constructed and successfully expressed in actinomycetes. We developed a marker-free expression method based on the combination of phage integration systems and site-specific recombinases. The Cre recombinase has been used in the deletion of huge genomic regions, including the phenalinolactone, monensin, and lipomycin biosynthetic gene clusters from Streptomyces sp. strain Tü6071, Streptomyces cinnamonensis A519, and Streptomyces aureofaciens Tü117, respectively. Finally, we also demonstrated the site-specific integration of plasmid and cosmid DNA into the chromosome of actinomycetes catalyzed by the Cre recombinase. We anticipate that the strategies presented here will be used extensively to study the genetics of actinomycetes.

Genome sequence of Streptomyces sp. strain Tü6071.

Streptomyces sp. Tü6071 is a soil-dwelling bacterium which has a highly active isoprenoid biosynthesis. Isoprenoids are important precursors for biopharmaceutical molecules such as antibiotics or anticancer agents, e.g., landomycin. Streptomyces sp. Tü6071 produces the industrially important terpene glycosides phenalinolactones, which have antibacterial activity against several Gram-positive bacteria. The availability of the genome sequence of Streptomyces sp. Tü6071 allows for understanding the biosynthesis of these pharmaceutical molecules and will facilitate rational genome modification to improve industrial use.