Research Area

Many clinically relevant drugs, such as penicillin, vancomycin, daptomycin or colistin are biocatalytically produced by microbes involving enzymes called non-ribosomal peptide synthetases (NRPS). These enzymes are able to produce molecules with structural features like cyclizations, acyl chains, D- or other non-proteinogenic amino acids or N-methylations, which confer those molecules favorable drug-like properties. Thus, natural products have historically been a rich source for drug discovery, accounting for around 30% of small-molecule drugs approved between 2000 and 2020.

In nature, the recombination of non-ribosomal peptide synthetase (NRPS) genes drives the evolution of new natural products with diverse bioactivities. Inspired by this process, it has been a longstanding goal to engineer NRPS enzymes in the laboratory to generate novel compounds with new biological activities. However, despite their seemingly modular architecture, NRPS enzymes remain notoriously difficult to reprogram in a predictable manner.

In our group, we aim to develop design principles and novel tools for predictable engineering of NRPS enzymes. Recently, we have introduce split inteins as a novel tool for NRPS engineering. Split inteins are small proteins that can reassemble and splice themselves out while joining their flanking protein sequences, enabling the expression of functional NRPS enzymes from separate fragments. This approach allowed the generation of a peptide library comprising more than 200 new-to-nature peptides in E. coli. Using such tools, we are creating large datasets to better understand module compatibility in engineered NRPS systems. Moreover, peptide libraries generated through NRPS engineering represent valuable sources for the discovery of new antibiotics.

Illustration of bacterial genomic pathways and chemical synthesis, showing engineered NRPS and new peptides creation. — **Figure 1. | NRPS engineering workflow for the production of novel non-ribosomal peptides in *E. coli*.** NRPS enzymes are involved in the biosynthesis of various natural products in bacteria. Here shown are the natural product bicornitin from *Xenorhabdus* *budapestensis*, ambactin from *Xenorhabdus indica* and GameXpeptide from *Photorhabdus laumondii*. Genetic recombination of NPRS genes from different organisms and heterologous expression allows the production of novel peptides in *E. coli*. Such new-to-nature peptides can then be screened for novel bioactivities.

**Figure 1. | NRPS engineering workflow for the production of novel non-ribosomal peptides in *E. coli*.** NRPS enzymes are involved in the biosynthesis of various natural products in bacteria. Here shown are the natural product bicornitin from *Xenorhabdus* *budapestensis*, ambactin from *Xenorhabdus indica* and GameXpeptide from *Photorhabdus laumondii*. Genetic recombination of NPRS genes from different organisms and heterologous expression allows the production of novel peptides in *E. coli*. Such new-to-nature peptides can then be screened for novel bioactivities.

Our group supervised the iGEM Team Marburg 2025 with their project “NRPieceS – Unlocking Nature’s Antibiotic Toolbox”. More information about this project can be found on their wiki page: https://2025.igem.wiki/marburg/.