Expanding the actinomycetes landscape for phosphonate natural products through genome mining

Phosphonate natural products (P-NPs) represent a unique and underexplored class of bioactive compounds with significant pharmaceutical and biotechnological potential. Many novel P-NPs with promising bioactivities were identified in recent years by genome mining of actinomycetes. The DSMZ strain collection harbors more than 6000 actinobacterial strains including an increasing number of genome sequenced strains. In this study, 940 genome-sequenced actinomycetes from the DSMZ and University of Tübingen strain collections were screened for the presence of phosphonate biosynthetic gene clusters (P-BGCs) by searching for the conserved pepM gene. This effort led to the identification of 54 potential phosphonate producer strains. Subsequent bioassays with a phosphonate-sensitive E. coli test strain showed activity for 17 strains, and ³¹P NMR spectroscopic analysis of culture supernatants confirmed phosphonate production for 21 strains, including the rare actinomycete Kitasatospora fiedleri DSM 114396^T. The functionality of the unique K. fiedleri P-BGC was verified by pepM gene deletion, which abolished phosphonate production in K. fiedleri, whereas overexpression of a cluster-situated LuxR-like regulator improved phosphonate production. These findings highlight the P-NP biosynthetic potential of actinomycetes and pave the way for discovering novel bioactive phosphonates.