Biology subjects

Cuthbert, B. J.

Publications and source records attributed to Cuthbert, B. J..

4 recordsLinked to original sources

bioRxiv2026

A glyoxal sensing Pseudomonas aeruginosa transcription factor enables lung infection

Aldehydes are a class of normally unwanted toxic electrophilic compounds that mainly arise from oxidation of glucose, lipids or DNA. However, it has recently come to light that they can also be weaponized by professional phagocytes to kill engulfed bacteria. How microbes subvert these assaults remains largely enigmatic. Here we describe the function, atomic structure and mechanism of the first bacterial transcription factor able to directly sense the dicarbonyl glyoxal (GO), which we aptly named the Glyoxal Regulator (GloR, from Pseudomonas aeruginosa PAO1), We show that GloR directly senses GO through a reversible cysteine modification that results in its binding to a conserved DNA regulatory motif (a glo box), which then triggers a transcriptional activation of a defined set of genes to help counter GO toxicity and enable acute lung infection. Despite substantial evolutionary divergence, when unmodified gloR and a glo box-regulated reporter were transferred into E. coli, a strikingly tight GO-specific regulation was maintained, suggesting this system could be readily transferred between unrelated microbial species. As homologs of GloR were identified in diverse bacterial species we anticipate its use to be widespread in both pathogens and environmental bacteria. Taken together, we present the first bona fide bacterial aldehyde regulator which senses host GO to enable survival during infection.

microbiology↗

bioRxiv2026

Redox regulated auto-processing controls delivery of an antibacterial cysteine peptidase toxin

Contact-dependent growth inhibition (CDI) is a mechanism of inter-bacterial competition mediated by CdiA effectors, which deliver polymorphic C-terminal toxins (CT) into neighboring competitors. StbD from Citrobacter rodentium DBS100 is an unusual CdiA-like protein that carries a C-terminal cysteine peptidase toxin. Crystallography reveals that StbD-CT is composed of an N-terminal cytoplasm-entry domain connected to a C39 family peptidase by a flexible linker. The entry domain hijacks membrane-embedded YajC for translocation into the target-cell cytosol where the peptidase inactivates type II topoisomerases. Intoxication leads to a loss of DNA super-helicity, impaired chromosome segregation and cell filamentation. In addition to cleaving topoisomerases, StbD-CT exhibits auto-proteolytic processing under reducing conditions, and this activity is required for target cell intoxication. We propose that StbD-CT remains tethered to the cell periphery via interactions with YajC after delivery. Auto-processing releases the peptidase, enabling the domain to penetrate into the cell interior where it cleaves nucleoid-associated topoisomerases. Together, these findings identify a proteolytic effector that deactivates type II topoisomerases and reveal a redox regulatory strategy that coordinates toxin activation with intercellular delivery.

microbiology↗

bioRxiv2025

Bacteria deliver a microtubule-binding protein into mammalian cells to promote colonization

Pathogenic Bordetella bacteria infect the ciliated respiratory epithelia of mammalian and avian hosts. Several bacterial proteins mediate host cell adhesion, but filamentous hemagglutinin (FhaB) is a principal adhesin because mutants lacking this protein exhibit profound colonization defects. Here, we show that FhaB carries a C-terminal microtubule-binding domain (FhaB-CT), which is translocated into the host-cell cytoplasm to promote bacterial colonization. Cryogenic electron microscopy of microtubule-bound FhaB-CT shows that the domain binds primarily to -tubulin through a network of polar interactions. Live-cell microscopy of infected tracheal explants reveals that FhaB-CT delivery is required for Bordetella to occupy a niche at the base of cilia on airway epithelia. Finally, we demonstrate that the microtubule-binding domain is required for long-term colonization of the mouse nasal cavity by B. pertussis. These observations suggest that the FhaB-CT domain is delivered into motile cilia, where it interacts with axonemal microtubules. We propose that Bordetella initially adhere to the tips of cilia, then deploy multiple FhaB adhesin molecules to migrate to the base of the cilial forest. This mechanism enables Bordetella to resist removal by the mucociliary escalator that clears the respiratory tract of microbes and debris.

microbiology↗

bioRxiv2020

Structural and molecular dynamics of Mycobacterium tuberculosis malic enzyme, a potential anti-TB drug target

Tuberculosis (TB) is the most lethal bacterial infectious disease worldwide. It is notoriously difficult to treat, requiring a cocktail of antibiotics administered over many months. The dense, waxy outer membrane of the TB-causing agent, Mycobacterium tuberculosis (Mtb), acts as a formidable barrier against uptake of antibiotics. Subsequently, enzymes involved in maintaining the integrity of the Mtb cell wall are promising drug targets. Recently, we demonstrated that Mtb lacking malic enzyme (MEZ) has altered cell wall lipid composition and attenuated uptake by macrophages. These results suggest that MEZ provides the required reducing power for lipid biosynthesis. Here, we present the X-ray crystal structure of MEZ to 3.6 [A] resolution and compare it with known structures of prokaryotic and eukaryotic malic enzymes. We use biochemical assays to determine its oligomeric state and to evaluate the effects of pH and allosteric regulators on its kinetics and thermal stability. To assess the interactions between MEZ and its substrate malate and cofactors, Mn2+ and NAD(P)+, we ran a series of molecular dynamics (MD) simulations. First, the MD analysis corroborates our empirical observations that MEZ is unusually disordered, which persists even with the addition of substrate and cofactors. Second, the MD simulations reveal that MEZ subunits alternate between open and closed states and that MEZ can stably bind its NAD(P)+ cofactor in multiple conformations, including an inactive, compact NAD+ form. Together the structure of MEZ and insights from its dynamics can be harnessed to inform the design of MEZ inhibitors that target Mtb.

biochemistry↗