“Erasing” Mutation Acquisition

Few examples highlight the implications of a dynamic genome more clearly than the challenge of antibiotic resistance. Underlying the pessimistic view permeating antimicrobial discovery is the belief that antibiotic resistance is inevitable. Our lab has suggested a new way of thinking of this challenge: Rather than accepting this tenet, can we uncover the mechanisms underlying bacterial evolution and use those insights to prevent antibiotic evasion?

A key pathway in acquired antibiotic resistance is the bacterial DNA damage response, also known as the SOS response. Compelling evidence from our lab (mSphere, 2016) and others has shown that inhibition of the SOS response slows the evolution of resistance, particularly with DNA damaging antibiotics such as fluoroquinolones. The SOS response is governed by the interaction between two proteins, RecA and LexA. In the setting of DNA damage, RecA forms long nucleoprotein filaments on ssDNA (termed ‘RecA*’). LexA is a repressor of SOS genes but can be induced to undergo self-proteolysis upon binding to RecA*. Recognizing the opportunity offered by targeting the SOS response, we devised the first high-throughput screen to discover inhibitors of RecA*-dependent LexA cleavage (ACS Infect Dis, 2018). We also developed a series of chemical biology tools to dissect each step in the process of RecA* formation, LexA binding, and LexA cleavage that set the stage for inhibitor optimization. Although cell permeation and potency remain bottlenecks, we have shown that our lead inhibitors can slow the rate of mutagenesis, offering a solid foundation for our novel approach.

We recognized that rational efforts to block SOS activation could be fueled by a molecular understanding of the LexA-RecA* complex; however, this structure had remained refractory to atomistic resolution despite decades of prior effort. In recent work using CryoEM approaches to resolve the heterogeneity resulting from LexA engagement with the RecA nucleoprotein helical filament, we reported the complete structure of the SOS activation complex (Nat Struct Mol Biol, 2024). This structure revealed an unexpected allosteric lock-and-key mechanism that offers immediate new avenues for small-molecule discovery.

Motivated by our prior drug discovery efforts and informed by structure and our mechanistic probes, with support from NIGMS, the Burroughs Wellcome Fund, and other sources, the vision for this part of our research program is to ultimately develop a new class of small-molecule SOS inhibitors. Akin to b-lactam/b-lactamase inhibitors, we aspire to combine these agents with DNA-damaging antibiotics (e.g., fluoroquinolones), thereby disabling the mechanisms engendering resistance (DISARMERs).