PT  - JOURNAL ARTICLE
AU  - Raphael Reher
AU  - Allegra T Aron
AU  - Pavla Fajtová
AU  - Paolo Stincone
AU  - Chenxi Liu
AU  - Ido Y Ben Shalom
AU  - Wout Bittremieux
AU  - Mingxun Wang
AU  - Marie L Matos-Hernandez
AU  - Kelsey L Alexander
AU  - Eduardo J Caro-Diaz
AU  - C Benjamin Naman
AU  - Chambers C. Hughes
AU  - Pieter C Dorrestein
AU  - Anthony J O’Donoghue
AU  - William H Gerwick
AU  - Daniel Petras
TI  - Native Metabolomics Identifies the Rivulariapeptolide Family of Protease Inhibitors
AID  - 10.1101/2021.09.03.458897
DP  - 2022 Jan 01
TA  - bioRxiv
PG  - 2021.09.03.458897
4099  - http://biorxiv.org/content/early/2022/02/06/2021.09.03.458897.short
4100  - http://biorxiv.org/content/early/2022/02/06/2021.09.03.458897.full
AB  - The identity and biological activity of most metabolites still remain unknown. A key bottleneck in the full exploration of this tremendous source of new structures and pharmaceutical activities is the compound purification needed for bioactivity assignments of individual compounds and downstream structure elucidation. To enable bioactivity-focused compound identification from complex mixtures, we developed a scalable native metabolomics approach that integrates non-targeted liquid chromatography tandem mass spectrometry, and simultaneous detection of protein binding via native mass spectrometry. While screening for new protease inhibitors from an environmental cyanobacteria community, native metabolomics revealed 30 cyclodepsipeptides as chymotrypsin binders. Mass spectrometry-guided purification then allowed for the full structure elucidation of four new specialized metabolites via tandem mass spectrometry, chemical derivatization, and nuclear magnetic resonance spectroscopy. Together with the evaluation of biological activities, our results identified the rivulariapeptolides as a family of serine protease inhibitors with nanomolar potency, highlighting native metabolomics as promising approach for drug discovery, chemical ecology, and chemical biology studies.Competing Interest StatementP.C.D. and W.H.G. are scientific advisors of Sirenas. P.C.D. is a scientific advisor of Galileo, Cybele, and scientific advisor and co-founder of Ometa Labs LLC and Enveda with approval by the UC San Diego. M.W. is a founder of Ometa Labs LLC.