Biological & mechanistic insights for targeting GroEL chaperonin systems for antibiotic development.

Wednesday, February 7, 2024


The escalating crisis of antimicrobial resistance (AMR) necessitates the exploration of innovative therapeutic candidates. Our research program is pioneering an approach that targets the GroEL chaperonin system, a 60 kDa heat shock protein that plays a vital role maintaining protein homeostasis within bacteria. It operates in an ATP-dependent manner through the assistance of GroES, a co-chaperone that caps the active GroEL rings and encapsulates unfolded proteins within the GroEL-GroES chamber, allowing them to fold to their functional forms isolated from the cellular milieu. Inhibiting this molecular machine induces a cascade effect that disrupts numerous essential cellular functions and leads to bacterial death. This novel strategy holds great promise in combating bacteria resistant to existing antibiotics as GroEL has not been previously targeted by other drugs. Our initial high-throughput screening yielded ~400 diverse GroEL inhibitors. Subsequent development has produced several series of analogs that exhibit potent and selective inhibition of E. coli and members of the ESKAPE bacteria. These lead inhibitors have demonstrated exceptional ability to rapidly eliminate bacteria, including those within biofilms that are traditionally resistant to antibiotics, while exhibiting low toxicity to human cells. Recent efforts have focused on identifying inhibitor mechanisms of action and target engagement in cells. Using E. coli and S. aureus engineered to express GFP, whose folding is facilitated by GroEL, we have established target engagement of lead inhibitors in bacteria. Through CryoEM and LC-MS/MS techniques, we have identified binding sites for several inhibitor series, uncovering distinct mechanisms that interfere with GroEL's critical functional transitions. This knowledge has enabled us to validate on-target effects by expressing GroEL constructs resistant to inhibitors in E. coli. Our research is now advancing into animal studies to assess the safety and antibiotic efficacy of lead inhibitors. The potential impact of our GroEL-targeting antibiotic strategy extends beyond combating antimicrobial resistance in the ESKAPE pathogens. We are expanding this approach to address critical global health threats such as Mycobacterium tuberculosis and Trypanosoma brucei, the causative agent of African Sleeping Sickness. Thus, our GroEL-targeting antibiotic strategy offers a novel and promising avenue to combat a wide range of infectious organisms and tackle the growing challenge of antimicrobial resistance.

Speaker Details

Dr. Steven M. Johnson

Indiana University

My research experiences have been a deliberate path towards acquiring the skills to lead cutting-edge,
multidisciplinary research programs. My industrial-to-academic training has been marked by a dedication
to innovative chemical biology research towards therapeutic development. My work has traversed the
preclinical drug discovery and development spectrum, providing me with a robust foundation from target
identification, validation, and high-throughput screening, to in vitro and in vivo hit-to-lead optimization.
My internships at Boehringer Ingelheim, where I synthesized drug candidates to target HCV NS3 protease
and HIV reverse transcriptase, sparked my enthusiasm for pursuing a career in biomedical research.
Towards this goal, I ventured into the protein misfolding and neurodegeneration fields during my graduate
studies with Professor Jeffery Kelly at The Scripps Research Institute, where I synthesized molecules to
study and selectively inhibit transthyretin amyloidogenesis. I continued investigating modulating protein
folding pathways during my postdoctoral studies with Professor Arthur Horwich at Scripps/Yale, where
we discovered inhibitors of the bacterial HSP60/10 chaperonin system (aka GroEL/ES) via highthroughput screening of a 700,000 compound library. At that point, I had solidified my desire to pursue
translational academic research geared at targeting protein homeostasis pathways of infectious organisms
as a novel antibiotic strategy. However, to gain further insights into infectious disease drug development,
I pursued a second postdoc with Professors Wesley Van Voorhis and Dustin Maly at the University of
Washington, where I developed inhibitors of calcium-dependent protein kinases from Toxoplasma gondii
and Cryptosporidium parvum as part of a multi-institute collaboration focusing on the pharmacological
optimization of antibiotics for toxoplasmosis and cryptosporidiosis. My current research at the Indiana
University School of Medicine expands on my interests and experiences in protein folding/misfolding to
investigate targeting protein homeostasis (proteostasis) pathways to develop new infectious disease and
anti-cancer therapeutics. We are leading a niche field investigating the potential of therapeutically
targeting GroEL/ES and HSP60/10 chaperonin systems. Through our studies, we have developed >1,000
GroEL/ES and HSP60/10 inhibitors, many of which have antibiotic effects against a range of bacteria and
parasites, and chemotherapeutic effects against colon, breast, and other cancer cells. Building from our
discoveries, I recently co-founded BioEL Inc. to translate our GroEL/ES-targeting antibacterial candidates
through pre-clinical development, and anticipate launching a second biotech company focused on
advancing our HSP60-targeting chemotherapeutics research program.