Method development for HPLC-MS



PhD Project: Identification and Characterization of Fungal RiPPs

Natural products as source for physical and mental well-being are accompanying mankind since ancient times [1]. Among the known classes of natural products, ribosomally synthesized and post-translationally modified peptides (RiPPs) constitute an attractive compound class for therapeutic developments, due to their bioactive properties (e.g. antimicrobial [2], antiviral [3], etc.). In contrast to the many described RiPP classes of bacterial origin, only eight classes of fungal RiPPs have been discovered so far [4] – a well of untapped potential. Despite the possibility to predict RiPP-precursor peptide sequences by genome mining tools, the numerous and diverse post-translational modifications arising during their maturation make it virtually impossible to predict the final structure.

Through this project, we aim to identify and characterize novel fungal RiPPs and elucidate the biosynthetic pathways with are involved in production of those. After identification of potential fungal RiPP precursor sequences, expression of the individual RiPP precursors is experimentally verified by transcriptome analysis and quantitative proteomics. For that, a comparative proteomics study between RiPP knock-out and overexpression fungal strains is conducted. It is then analysed with a special focus on the precursor-RiPP and the modifying enzymes in the RiPP gene cluster.

For identification of mature RiPPs, whose final structure is not known, quantitative untargeted metabolomics approaches are used, applying open modification search approaches or molecular networking tools for data evaluation. We envision that via molecular network analysis intermediate biosynthetic steps starting from the precursor can be tracked to and unravel the mature RiPP mass and biosynthetic RiPP pathways.


[1]          D. A. Dias, S. Urban, and U. Roessner, “A Historical overview of natural products in drug discovery,” Metabolites, vol. 2, no. 2. Molecular Diversity Preservation International, pp. 303–336, Apr. 16, 2012, doi: 10.3390/metabo2020303.

[2]          P. G. Arnison et al., “Ribosomally synthesized and post-translationally modified peptide natural products: Overview and recommendations for a universal nomenclature,” Nat. Prod. Rep., vol. 30, no. 1, pp. 108–160, 2013, doi: 10.1039/c2np20085f.

[3]          Y. Fu, A. H. Jaarsma, and O. P. Kuipers, “Antiviral activities and applications of ribosomally synthesized and post-translationally modified peptides (RiPPs),” Cellular and Molecular Life Sciences, vol. 78, no. 8. Springer, pp. 3921–3940, Feb. 02, 2021, doi: 10.1007/s00018-021-03759-0.

[4]          S. C. Kessler and Y.-H. Chooi, “Out for a RiPP: challenges and advances in genome mining of ribosomal peptides from fungi,” Nat. Prod. Rep., 2022, doi: 10.1039/d1np00048a.