# ClpC1-targeting peptide natural products differentially dysregulate the proteome of Mycobacterium tuberculosis

## TL;DR

This study shows how three natural peptides affect the protein system in tuberculosis bacteria differently, offering new ways to develop antibiotics.

## Contribution

The study reveals distinct proteomic effects of three ClpC1-targeting peptides in Mycobacterium tuberculosis.

## Key findings

- Ecumicin, ilamycin, and cyclomarin differentially dysregulate the Mycobacterium tuberculosis proteome.
- Ilamycin and ecumicin do not trigger the ClpC2 rescue mechanism activated by cyclomarin.
- Ecumicin interacts with the stress-response chaperone Hsp20.

## Abstract

Targeting the protein quality control system in Mycobacterium tuberculosis represents a promising and underexplored opportunity for antibiotic development. The ClpC1:ClpP1P2 protease is an essential component of the system that mediates both regulatory and stress-related protein degradation. Several non-ribosomal peptide natural products, including ecumicin, ilamycins (rufomycins) and cyclomarins, have been discovered that bind to the ClpC1 chaperone of the complex and exhibit potent antimycobacterial activity, leading to significant interest in the ClpC1:ClpP1P2 system as a bona fide target for the new tuberculosis drugs. In this study, we combine quantitative proteomics, bioinformatics, transcriptomics, CRISPRi knockdown, and targeted biochemical and biophysical assays to dissect the mechanisms of ecumicin, ilamycin and cyclomarin in clinically relevant Mycobacterium tuberculosis. Strikingly, despite exhibiting similar binding modes to ClpC1, each compound induces distinct effects on protein degradation. Notably, ilamycin and ecumicin do not trigger the ClpC2 rescue mechanism that mitigates cyclomarin-induced mycobacterial toxicity. In addition, we identify a novel interaction between ecumicin and stress-response chaperone Hsp20. The differential disruption of ClpC1 substrates, stress-response chaperones, and distinct reshaping of the Mycobacterium tuberculosis proteome by the three natural products, unveils new opportunities for the development of protein quality control-targeted antimycobacterials.

In this work, the authors reveal that three non-ribosomal peptides targeting a common protein quality system differentially alter the Mycobacterium tuberculosis proteome, providing new insight into the molecular mechanisms behind their antimycobacterial activity.