Class IIb Microcin MccM Interferes with Oxidative Phosphorylation in Escherichia coli

Dysbiosis
of the human gut microbiota is linked to numerous diseases.
Understanding the molecular mechanisms by which microbes interact
and compete with one another is required for developing successful
strategies to modulate the microbiome. The natural product Microcin
M (MccM) consists of a 77-residue bioactive peptide conjugated to
a siderophore and is a class II microcin involved in microbial competition
with an enigmatic mode-of-action. In this work, we investigated the
basis for MccM activity and leveraged bioinformatics to expand the
known chemical diversity of class II microcins. We applied automated
fast-flow solid phase peptide synthesis coupled with chemoenzymatic
chemistry to acquire MccM and demonstrated that its activity was bacteriostatic.
We then used our synthetic molecule to ascertain that catecholate
siderophore transporters in Escherichia coli K-12 are necessary for MccM import. Once inside the cell, we found
that MccM treatment decreased the levels of intracellular ATP and
interfered with gene expression. These effects were ameliorated in
genetic mutants lacking ATP synthase or in conditions that support
substrate-level phosphorylation. Further, we showed that MccM elevated
the levels of reactive oxygen species within the target cell. We propose
that MccM effects its bacteriostatic activity by decreasing the total
energy level of the cell through inhibition of oxidative phosphorylation.
Lastly, using genome mining, we bioinformatically identified 171 novel
putative class II microcins. Our investigation sheds light on the
natural processes involved in microbial competition and provides inspiration,
in the form of new molecules, for future therapeutic endeavors.