Odilorhabdin, a promising antibiotic candidate, has the potential to combat antibiotic-resistant bacteria. Nevertheless, the yield of the antibiotic in bacterial cultures remains insufficient. With the assistance of a technique known as NRPS engineering, researchers at Helmholtz Institute for Pharmaceutical Research Saarland (HIPS), in collaboration with the Max Planck Institute for Terrestrial Microbiology in Marburg, have elucidated the biosynthesis pathway and identified potential avenues for the future synthesis and biological production of the antibiotic. These findings contribute to the accelerated development of effective antibiotics in response to the growing prevalence of bacterial resistance.
The peptide antibiotic odilorhabdin, which was only recently identified, has been demonstrated to be effective against a wide range of bacterial species, including those that have developed resistance to other antibiotics. This renders odilorhabdin a promising candidate for a novel class of prospective pharmaceutical agents. Nevertheless, the route from potential active ingredient to drug is not straightforward. As natural producers, bacteria often exhibit low levels of productivity, resulting in the production of the desired substance in only very small quantities. Similarly, the yield of odilorhabdin produced by microorganisms remains insufficient, so that an extensive analysis of its production has so far not seemed feasible.
Recently, researchers from Prof. Rolf Müller's Department of Microbial Natural Products at HIPS, in collaboration with researchers led by Prof. Helge Bode from the Max Planck Institute for Terrestrial Microbiology in Marburg and the French biotechnology company Nosopharm, have achieved a comprehensive understanding of the antibiotic's formation.
The key to this was a technique known as NRPS engineering, which involves modifying genetic blueprints for non-ribosomal peptide synthetases (NRPS) – giant bacterial enzymes found in bacteria and fungi that produce some of the most important clinically used antibiotics, including penicillin and other therapeutic agents. Enzymes with new capabilities are created, which in turn produce modified antibiotics.
Odilorhabdin, like many other so-called peptide antibiotics, has so far been challenging to identify using these techniques. “The genetic blueprint of odilorhabdin, the so-called NRPS gene cluster, was subjected to detailed analysis and modification in the original producer Xenorhabdus nematophila. We then transferred the new blueprint into a microbial producer that can be easily cultivated in the laboratory," states Dr. Carsten Seyfert, scientist in the Department of Microbial Natural Products at HIPS. "Despite the initial suboptimal production in the bacterium E. coli due to the inherent toxicity of odilorhabdin, we devised an innovative approach to successfully investigate the gene cluster in collaboration with Helge Bode's team."
The trick: the researchers took the antibiotic piece by piece and coupled its parts to known biosynthesis genes. This allowed them to utilize the new functional hybrids to investigate the specific roles of each section. The hybrids are non-toxic variations of the gene cluster, thereby enabling the researchers to investigate all relevant parts of the biosynthesis.
“This breakthrough demonstrates the important role that interdisciplinary collaboration plays in the field of antibiotic research. Our ability to fully decipher the biosynthesis of odilorhabdin is a significant step towards its biotechnological production. This should allow us to produce our drug candidate cost-effectively and in large quantities - a crucial step on the way to a potential application,” states Prof. Rolf Müller, Managing Director of the HIPS and Head of the Department of Microbial Natural Products.
Additionally, the team demonstrated that odilorhabdin is initially produced as a so-called pro-drug. An inactive precursor can only be cleaved into the active antibiotic after it has been transported out of the cell. This facilitates the future production of numerous variants that could enable a broad spectrum of antimicrobial effects as representatives of this novel class of antibiotics through the use of NPRS engineering.
Original Publication
Leonard Präve, Carsten E. Seyfert, Kenan A. J. Bozhüyük, Emilie Racine, Rolf Müller, Helge B. Bode. Investigation of the Odilorhabdin Biosynthetic Gene Cluster Using NRPS Engineering. L. Präve, C. E. Seyfert, K. A. J. Bozhüyük, E. Racine, R. Müller, H. B. Bode, Angew. Chem. Int. Ed. 2024, 63, e202406389. DOI: 10.1002/anie.202406389
