Biomimetic synthesis, biosynthetic pathway engineering and structure activities studies of unique glycosolyated macrolactams

• Applicant: Professorin Dr. Christine Beemelmanns
• Subject Area: Biological and Biomimetic Chemistry ; Organic Molecular Chemistry - Synthesis and Characterisation
• Term: from 2017 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 391056593

• Final Report Year: 2023

Final Report Abstract

With the global spread of multiresistant pathogenic bacterial strains, such as methicillin resistant Staphylococcus aureus (MRSA) and vancomycin resistant Enterococci (VRE), the demand for new classes of antibiotics, including those from natural sources, has been steadily increasing again. The genome and metabolome-driven analysis of microbial symbionts yields not only a deeper understanding of the evolution of symbiotic systems, but also deeper insights into the diversity of biosynthetic pathways and accesses to novel natural products. This project aimed to investigate bacterial symbionts of termites, with a particular focus on those isolates that are capable of producing antibiotic macrolactams. In this project, we capitalized on recent advances in DNA sequencing technologies and performed a comparative genome-mining driven approach of bacterial symbionts to evaluate their secondary metabolite repertoire and the diversity of biosynthetic pathways encoding the production of macrolactams. In frame of this study, we were able to revise the originally proposed biosynthetic pathway of macrotermycins produced by Amycolatopsis sp. M39 based on newly acquired genome sequencing data, and identified a new macrolactam pathway (pac, asa) in a new Amycolatopsis isolate. We then made use of a dual promotor vector to build up a heterologous production system in a Streptomycetes host enabling us now to engineer the biosynthetic pathway. Based on these studies, we were able to verify the asa macrolactam biosynthetic pathway. Using a coculture set-ups, we also evaluated means by which macrolactam biosynthetic gene clusters are stimulated in wild types and were able to isolate four macrotermycin derivatives from strain M39 and circomycin A from Amycolatopsis sp. PS_44_ISF1. Meanwhile, our collaborators (ANR) synthesized key fragments towards the core structure, which will in future assist in the total synthesis and absolute structure verification of macrotermycins.

Publications

• Actinomadura rubteroloni sp. nov. and Actinomadura macrotermitis sp. nov., isolated from the gut of the fungus growing-termite Macrotermes natalensis. International Journal of Systematic and Evolutionary Microbiology, 70(10), 5255-5262.
  Benndorf, René; Martin, Karin; Küfner, Michelle; de Beer, Z. Wilhelm; Vollmers, John; Kaster, Anne-Kristin & Beemelmanns, Christine
  
• Nocardia macrotermitis sp. nov. and Nocardia aurantia sp. nov., isolated from the gut of the fungus-growing termite Macrotermes natalensis. International Journal of Systematic and Evolutionary Microbiology, 70(10), 5226-5234.
  Benndorf, René; Schwitalla, Jan W.; Martin, Karin; de Beer, Z. Wilhelm; Vollmers, John; Kaster, Anne-Kristin; Poulsen, Michael & Beemelmanns, Christine
  
• Polyhalogenation of Isoflavonoids by the Termite-Associated Actinomadura sp. RB99. Journal of Natural Products, 83(10), 3102-3110.
  Rak, Lee Seoung; Schalk, Felix; Schwitalla, Jan W.; Benndorf, René; Vollmers, John; Kaster, Anne-Kristin; de Beer, Z. Wilhelm; Park, Minji; Ahn, Mi-Jeong; Jung, Won Hee; Beemelmanns, Christine & Kim, Ki Hyun
  
• Stereoselective Construction of (E,Z)‐1,3‐Dienes and Its Application in Natural Product Synthesis. Advanced Synthesis & Catalysis, 362(24), 5532-5575.
  Hubert, Pierre; Seibel, Elena; Beemelmanns, Christine; de Campagne, Jean‐Marc & Figueiredo, Renata Marcia
  
• Streptomyces smaragdinus sp. nov., isolated from the gut of the fungus growing-termite Macrotermes natalensis. International Journal of Systematic and Evolutionary Microbiology, 70(11), 5806-5811.
  Schwitalla, Jan W.; Benndorf, René; Martin, Karin; Vollmers, John; Kaster, Anne-Kristin; de Beer, Z. Wilhelm; Poulsen, Michael & Beemelmanns, Christine
  
• Targeted Discovery of Tetrapeptides and Cyclic Polyketide‐Peptide Hybrids from a Fungal Antagonist of Farming Termites. ChemBioChem, 21(20), 2991-2996.
  Schalk, Felix; Um, Soohyun; Guo, Huijuan; Kreuzenbeck, Nina B.; Görls, Helmar; de Beer, Z. Wilhelm & Beemelmanns, Christine
  
• Comparative Genomics Reveals Prophylactic and Catabolic Capabilities of Actinobacteria within the Fungus-Farming Termite Symbiosis. mSphere, 6(2).
  Murphy, Robert; Benndorf, René; de Beer, Z. Wilhelm; Vollmers, John; Kaster, Anne-Kristin; Beemelmanns, Christine & Poulsen, Michael
  
• Female‐biased sex allocation and lack of inbreeding avoidance in Cubitermes termites. Ecology and Evolution, 11(10), 5598-5605.
  Sinotte, Veronica M.; Conlon, Benjamin H.; Seibel, Elena; Schwitalla, Jan W.; de Beer, Z. Wilhelm; Poulsen, Michael & Bos, Nick
  
• Targeted Isolation of Saalfelduracin B–D from Amycolatopsis saalfeldensis Using LC-MS/MS-Based Molecular Networking. Journal of Natural Products, 84(4), 1002-1011.
  Um, Soohyun; Seibel, Elena; Schalk, Felix; Balluff, Sven & Beemelmanns, Christine
  
• Comparative Genomic and Metabolomic Analysis of Termitomyces Species Provides Insights into the Terpenome of the Fungal Cultivar and the Characteristic Odor of the Fungus Garden of Macrotermes natalensis Termites. mSystems, 7(1).
  Kreuzenbeck, Nina B.; Seibel, Elena; Schwitalla, Jan W.; Fricke, Janis; Conlon, Benjamin H.; Schmidt, Suzanne; Hammerbacher, Almuth; Köllner, Tobias G.; Poulsen, Michael; Hoffmeister, Dirk & Beemelmanns, Christine
  
• Insights into the Metabolomic Capacity of Podaxis and Isolation of Podaxisterols A–D, Ergosterol Derivatives Carrying Nitrosyl Cyanide-Derived Modifications. Journal of Natural Products, 85(9), 2159-2167.
  Guo, Huijuan; Daniel, Jan-Martin; Seibel, Elena; Burkhardt, Immo; Conlon, Benjamin H.; Görls, Helmar; Vassão, Daniel Giddings; Dickschat, Jeroen S.; Poulsen, Michael & Beemelmanns, Christine
  
• Adaptations of Pseudoxylaria towards a comb-associated lifestyle in fungus-farming termite colonies. The ISME Journal, 17(5), 733-747.
  Fricke, Janis; Schalk, Felix; Kreuzenbeck, Nina B.; Seibel, Elena; Hoffmann, Judith; Dittmann, Georg; Conlon, Benjamin H.; Guo, Huijuan; Wilhelm, de Beer Z.; Vassão, Daniel Giddings; Gleixner, Gerd; Poulsen, Michael & Beemelmanns, Christine