Rufomycin Targets ClpC1 Proteolysis in Mycobacterium tuberculosis and M. abscessus

Abstract

ClpC1 is an emerging new target for the treatment of <i>Mycobacterium tuberculosis</i> infections, and several cyclic peptides (ecumicin, cyclomarin A, and lassomycin) are known to act on this target. This study identified another group of peptides, the rufomycins (RUFs), as bactericidal to <i>M. tuberculosis</i> through the inhibition of ClpC1 and subsequent modulation of protein degradation of intracellular proteins. Rufomycin I (RUFI) was found to be a potent and selective lead compound for both <i>M. tuberculosis</i> (MIC, 0.02 μM) and <i>Mycobacterium abscessus</i> (MIC, 0.4 μM). Spontaneously generated mutants resistant to RUFI involved seven unique single nucleotide polymorphism (SNP) mutations at three distinct codons within the <i>N</i>-terminal domain of <i>clpC1</i> (V13, H77, and F80). RUFI also significantly decreased the proteolytic capabilities of the ClpC1/P1/P2 complex to degrade casein, while having no significant effect on the ATPase activity of ClpC1. This represents a marked difference from ecumicin, which inhibits ClpC1 proteolysis but stimulates the ATPase activity, thereby providing evidence that although these peptides share ClpC1 as a macromolecular target, their downstream effects are distinct, likely due to differences in binding.