Biology subjects

Kany, A. M.

Publications and source records attributed to Kany, A. M..

6 recordsLinked to original sources

bioRxiv2026

Pathogen-driven reactivation of metabolite prodrugs defines nitroxoline's iron-deprivation antibiotic activity

The rise of antimicrobial resistance warrants renewed attention to established but overlooked antibiotics such as nitroxoline (NTX). Here, we systematically dissect NTXs mode of action and investigate the contribution of its first-pass metabolites, NTX-sulphate and NTX-glucuronide. We identified metallophore-mediated cellular iron deprivation as the principal antibacterial mechanism of NTX, characterized by induction of iron acquisition pathways and Fe-S cluster proteins, and concomitant loss of protein-bound iron. In contrast, NTX metabolites were biologically inactive and lacked metal-chelating properties. Ex vivo assays demonstrated that clinically relevant uropathogens, including Escherichia coli and Klebsiella pneumoniae, efficiently reconvert these metabolites into active NTX in human urine. Together, our findings establish a mechanistic framework linking NTX antibacterial activity, host detoxification, and pathogen-dependent metabolite reactivation, and providing a molecular explanation for NTXs enduring therapeutic potential and favourable safety profile.

microbiology↗

bioRxiv2026

Generative AI designs functional thiolation domains for reprogramming non-ribosomal peptide synthetases

Large language models and generative protein design promise to accelerate biotechnology, but it remains unclear whether they can engineer dynamic megasynth(et)ases whose activity depends on transient, context-specific domain interfaces. Non-ribosomal peptide synthetases (NRPSs) are an especially demanding target, yet a high-value one because they produce many clinically important natural products and offer a route to analogs that are often difficult or impractical to access by chemical synthesis. Here we integrate pretrained generative models (ESM3, ProteinMPNN and EvoDiff) with design-build-test-learn cycles and data-guided prioritization to generate 76 de novo thiolation (T) domains. We built and tested 578 recombinant NRPS variants in vivo spanning minimal, full-length and hybrid assembly lines. AI-designed T-domains supported product formation across architectures, enabled catalytically active hybrids at recombined junctions and increased yields by up to [~]3-fold relative to NRPSs carrying the native T-domain. A representative design showed improved soluble expression, refolding, and a 12 {degrees}C higher melting temperature, while molecular dynamics simulations indicated preserved global stability but reshaped, state-dependent interdomain contact networks. Together, these results establish generative design as an effective route to context-conditioned optimization and reprogramming of biosynthetic assembly lines.

synthetic biology↗

bioRxiv2026

Structure Elucidation, Biosynthesis and Biological Evaluation of Neosorangicin A, a Member of the Sorangicin Family

Antimicrobial resistance represents an escalating global health crisis, with drug-resistant infections predicted to cause up to 10 million deaths annually by 2050, underscoring the urgent need for novel antibiotics. Natural products play a crucial role in the discovery and development of antibiotics, with myxobacteria emerging as a particularly promising source due to their ability to produce structurally diverse and bioactive compounds. One prominent example of antibiotics from myxobacteria are the sorangicins, potent inhibitors of the bacterial RNA polymerase (RNAP). Here, we report the isolation of two unprecedented compounds, neosorangicin A (1) and neosorangioside A (2), from Sorangium cellulosum strain Soce439, elucidated their molecular structures, thereby revealing significant structural variation in comparison to sorangicin, and describe their biosynthetic pathway. Neosorangicin A (1) exhibited strong activity against various Gram-positive bacteria, with enhanced potency on intracellular Staphylococcus aureus. In a murine wound infection model, a head-to-head comparison of neosorangicin A (1) and sorangicin A (3) provided useful insights into how the altered physicochemical properties, arising from the shortened side chain and the lack of the free carboxylic acid of neosorangicin A, influence the in vivo efficacy of sorangicin derivatives.

microbiology↗

bioRxiv2025

Tracheal tuft cell-released leukotrienes promote antibacterial immune responses

Tuft cells act as crucial sentinels in the airways that detect bacterial metabolites. In response, tuft cells release signaling molecules that trigger immune responses essential for clearing the infection. The molecular mechanisms driving immune cell activation following tuft cell stimulation in pneumonia are still not fully understood. Here, we identify tuft cells as the primary source of proinflammatory leukotrienes (LTs), which are released in the presence of pathogenic bacteria in the airways. We show that tracheal tuft cells discriminate pathogenic from non-pathogenic bacteria by sensing adenosine triphosphate (ATP) released from pathogens such as Pseudomonas aeruginosa and Rodentibacter pneumotropicus within the first 4 h of invasion, and recruit neutrophils and macrophages to the trachea and alveolar spaces. Taste signaling through the chemosensory transient receptor potential cation channel subfamily M member 5 (Trpm5) channel was essential for tuft cell activation and LT release. Mice lacking Trpm5 were not capable of detecting bacteria-released ATP and became colonized upon R. pneumotropicus infection. In contrast, Trpm5+/+ mice cleared the pathogen. We uncover a critical tuft cell-dependent sensing mechanism in pneumonia and establish tracheal tuft cells as both detectors of bacterial extracellular ATP and triggers of acute innate immune responses.

immunology↗

bioRxiv2024

Cystobactamid off-target profiling reveals favorable safety, superoxide reduction, and SCARB1 inhibition in eukaryotes

Antimicrobial resistance (AMR) poses a fundamental global threat, necessitating new strategies for effective therapies. Cystobactamids (CYS), a class of antibacterial agents targeting bacterial gyrase and topoisomerase IV, represent a non-traditional chemical scaffold with broad-spectrum activity. For toxicological de-risking, we performed a comprehensive profiling on eukaryotic cells, focusing on cytotoxicity, genotoxicity, and mitochondrial toxicity, demonstrating cellular safety and superoxide scavenging properties. Studies in zebrafish embryos assessed developmental, cardiovascular, and hepatic toxicity, indicating a favorable in vivo safety profile. Metabolism studies revealed glucuronidation and amide bond hydrolysis as key pathways, whereby CYS metabolic stability substantially improved by cobicistat co-treatment. Affinity-based protein profiling identified the cholesterol- and HCV-receptor scavenger receptor class B member 1 (SCARB1) as a primary eukaryotic off-target protein, with cystobactamids shown to inhibit SCARB1s function, preventing hepatitis C virus pseudoparticle entry into cells. These findings suggest a high therapeutic potential for cystobactamids and highlight SCARB1 as a primary eukaryotic target.

pharmacology and toxicology↗

bioRxiv2024

Thermally activated antibiotic production by probiotic bacteria for pathogen elimination

Peptide drugs have seen rapid advancement in biopharmaceutical development, with over 80 candidates approved globally. Despite their therapeutic potential, the clinical translation of peptide drugs is hampered by challenges in production yields and stability. Engineered bacterial therapeutics is a unique approach being explored to overcome these issues by using bacteria to produce and deliver therapeutic compounds at the body site of use. A key advantage of this technology is the possibility to control drug delivery within the body in real time using genetic switches. However, the performance of such genetic switches suffers when used to control drugs that require post-translational modifications or are toxic to the host. In this study, these challenges were experienced when attempting to establish a thermal switch for the production of a ribosomally synthesized and post-translationally modified peptide antibiotic, darobactin, in probiotic E. coli. These challenges were overcome by developing a thermo-amplifier circuit that combined the thermal-switch with a T7 RNA Polymerase and its promoter that overcame limitations imposed by the host transcriptional machinery due to its orthogonality to it. This circuit enabled production of pathogen-inhibitory levels of darobactin at 40{degrees}C while maintaining leakiness below the detection limit at 37{degrees}C. More impressively, the thermo-amplifier circuit sustained production beyond the thermal induction duration. Thus, raised temperature for 2 h was sufficient for the bacteria to produce pathogen-inhibitory levels of darobactin even in the physiologically relevant simulated conditions of the intestines that include bile salts and low nutrient levels. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=138 SRC="FIGDIR/small/579303v2_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@1cdb311org.highwire.dtl.DTLVardef@1a7523aorg.highwire.dtl.DTLVardef@77b68org.highwire.dtl.DTLVardef@136f52b_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗