Exploitation of genome mining tools for accelerated natural product discovery

Streptomycetes are rich sources of new bioactive natural products. Their genomes typically contain over twenty biosynthetic gene clusters encoding for secondary metabolite pathways [1]. One of those are nonribosomally produced peptides (NRP). They represent a structurally diverse family of natural products with a broad spectrum of biological activities, e. g. antibiotic, anti-cancer and immunosupressive [2], [3]. Clusters encoding for nonribosomal peptide synthetases (NRPS) can be easily identified by genome mining, i. e. screening and identification of these clusters employing bioinformatics [4].

Aim of the project is to identify novel gene clusters by genome mining and to isolate the predicted products of these clusters as well as to evaluate their bioactivity. An interesting candidate is Streptomyces fragilis which is predicted to produce an NRP by two NRPS [Fig. 1].

In order to isolate the predicted NRP, the substrate specificity of the adenylation domains of the two NRPS were determined by cloning, overexpression and γ-¹⁸O₄-ATP-pyrophosphate exchange assay [5]. l-Valine was found to be the predominant substrate for six out of nine tested adenylation domains. To identify a valine-rich natural product produced by S. fragilis, ¹⁴C-valine was fed to a 25 ml culture. Culture extracts were analyzed by radio-HPLC and radio-TLC for radioactive compounds. One radioactive product was detected. Its identification is in process.

Taken together, we have identified a cluster encoding for two NRPS with unusual substrate specificity which predicts the ability of this two NRPS to produce a new secondary metabolite.

Fig. 1 A gene cluster encoding two NRPS, which are predicted to produce an interesting NRP [4].

• References

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