Biotechnological production optimization of argyrins - a potent immunomodulatory natural product class

Argyrins represent a family of cyclic octapeptides exhibiting promising immunomodulatory activity via inhibiting mitochondrial protein synthesis, which leads to reduced IL-17 production by the T-helper 17 cells. Argyrins are formed by a non-ribosomal peptide synthetase (NRPS), originating from the myxobacterial producer strains Archangium gephyra Ar8082 and Cystobacter sp. SBCb004. In this work, a previously established heterologous production platform was employed to provide evidence of direct D-configured amino acid incorporation by the argyrin assembly line. An adenylation domain of the argyrin NRPS was characterized and shown to have a high preference for D-configured amino acids. Eight novel argyrin derivatives were generated via biosynthetic engineering of the heterologous production system. The system was also optimized to enable formation of methylated argyrin C and D derivatives with improved immunosuppressive activity compared with their unmethylated counterparts. Furthermore, the optimization of cultivation conditions allowed exclusive production of one major derivative at a time, drastically improving the purification process. Importantly, engineering of transcription and translation initiation resulted in a substantially improved production titre reaching 350-400 mg l^-1 . The optimized system presented herein thus provides a versatile platform for production of this promising class of immunosuppressants at a scale that should provide sufficient supply for upcoming pre-clinical development.