Biology subjects

Voyvodic, P. L.

Publications and source records attributed to Voyvodic, P. L..

6 recordsLinked to original sources

bioRxiv2026

Single-Cell Electrophysiology Reveals Verapamil's Disruption of Bacterial Membrane Energetics

Verapamil, a clinically used calcium channel blocker, enhances the activity of several tuberculosis antibiotics, but its mechanism of action and physiological effects on bacteria remain unresolved. A central debate concerns whether verapamil primarily inhibits efflux pumps or disrupts membrane energetics. Here, we use Escherichia coli as a model system to quantify single-cell and population-level physiological responses to verapamil with high temporal resolution. Real-time measurements of the rotational speed of individual flagellar motors, a single-cell proxy for the proton motive force (PMF), reveal a heterogeneous response to verapamil: treated cells exhibit either a dose-dependent gradual decrease in PMF, or a rapid collapse of PMF. Although loss of the outer-membrane efflux channel TolC increases growth inhibition by verapamil, it does not alter the rapid PMF disruptions observed at the single-cell level, suggesting that efflux contributes to long-term susceptibility but not to the initial PMF disruption. Independent assays of population-level motility, pH, and membrane-integrity suggest that verapamil may selectively dissipate the electrical component of PMF while leaving intracellular pH largely unchanged. A minimal electrical circuit model captures both steady-state and dynamic behavior. Together, these findings demonstrate that verapamil rapidly and reversibly perturbs bacterial membrane energetics through a mechanism distinct from classical protonophores, helping to reconcile conflicting interpretations of its activity and clarifying how membrane effects may interact with efflux inhibition during antibiotic potentiation.

biophysics↗

bioRxiv2022

PeroxiHUB: a modular cell-free biosensing platform using H2O2 as signal integrator

Cell-free systems have great potential for delivering robust, cheap, and field-deployable biosensors. Many cell-free biosensors rely on transcription factors responding to small molecules, but their discovery and implementation still remain challenging. Here we report the engineering of PeroxiHUB, an optimized H2O2-centered sensing platform supporting cell-free detection of different metabolites. H2O2 is a central metabolite and a by-product of numerous enzymatic reactions. PeroxiHUB uses enzymatic transducers to convert metabolites of interest into H2O2, enabling rapid reprogramming of sensor specificity using alternative transducers. We first screen several transcription factors and optimize OxyR for the transcriptional response to H2O2 in cell-free, highlighting the need for pre-incubation steps to obtain suitable signal-to-noise ratios. We then demonstrate modular detection of metabolites of clinical interest - lactate, sarcosine, and choline - using different transducers mined via a custom retro-synthesis workflow publicly available on the SynBioCAD Galaxy portal. We find that expressing the transducer during the pre-incubation step is crucial for optimal sensor operation. Finally, we show that different reporters can be connected to PeroxiHUB, providing high adaptability for various applications. Given the wide range of enzymatic reactions producing H2O2, the PeroxiHUB platform will support cell-free detection of a large number of metabolites in a modular and scalable fashion.

synthetic biology↗

bioRxiv2021

A synthetic receptor platform enables rapid and portable monitoring of liver dysfunction via engineered bacteria.

Bacterial biosensors, or bactosensors, are promising field-deployable agents for medical and environmental diagnostics. However, the lack of scalable frameworks to systematically program ligand detection limits their applications. Here we present a synthetic receptor platform, termed EMeRALD (Engineered Modularized Receptors Activated via Ligand-induced Dimerization) which supports the modular assembly of sensing modules onto a high-performance, generic signaling scaffold controlling gene expression in E. coli. We applied EMeRALD to detect bile salts, a biomarker of liver dysfunction, by repurposing sensing modules from enteropathogenic Vibrio species. We improved the sensitivity and lowered the limit-of-detection of the sensing module by directed evolution. We then engineered a colorimetric bactosensor detecting pathological bile salt levels in serum from patients having undergone liver transplant, providing an output detectable by the naked-eye. The EMeRALD technology enables functional exploration of natural sensing modules and rapid engineering of synthetic receptors for diagnostics, environmental monitoring, and control of therapeutic microbes.

synthetic biology↗

bioRxiv2020

Mechanical Tension in Syndecan-1 is Regulated by Extracellular Mechanical Cues and Fluidic Shear Stress

The endothelium plays a central role in regulating vascular homeostasis and is key in determining the response to materials implanted in the vascular system. Endothelial cells are uniquely sensitive to biophysical cues from applied forces and their local cellular microenvironment. The glycocalyx is a layer of proteoglycans, glycoproteins and glycosaminoglycans that lines the luminal surface of the vascular endothelium, interacting directly with the components of the blood and the forces of blood flow. In this work, we examined the changes in mechanical tension of syndecan-1, a cell surface proteoglycan that is an integral part of the glycocalyx, in response to substrate stiffness and fluidic shear stress. Our studies demonstrate that syndecan-1 is mechanically responsive to extracellular mechanical cues and alters its association with cytoskeletal and adhesion-related proteins in response to substrate stiffness and physiological flow.

bioengineering↗

bioRxiv2019

Metabolic Perceptrons for Neural Computing in Biological Systems

AbstractSynthetic biological circuits are promising tools for developing sophisticated systems for medical, industrial, and environmental applications. So far, circuit implementations commonly rely on gene expression regulation for information processing using digital logic. Here, we present a new approach for biological computation through metabolic circuits designed by computer-aided tools, implemented in both whole-cell and cell-free systems. We first combine metabolic transducers to build an analog adder, a device that sums up the concentrations of multiple input metabolites. Next, we build a weighted adder where the contributions of the different metabolites to the sum can be adjusted. Using a computational model trained on experimental data, we finally implement two four-input \"perceptrons\" for desired binary classification of metabolite combinations by applying model-predicted weights to the metabolic perceptron. The perceptron-mediated neural computing introduced here lays the groundwork for more advanced metabolic circuits for rapid and scalable multiplex sensing.

synthetic biology↗

bioRxiv2018

Plug-and-Play Metabolic Transducers Expand the Chemical Detection Space of Cell-Free Biosensors

Cell-free transcription-translation systems have great potential for biosensing, yet the range of detectable chemicals is limited. Here we provide a framework to expand the range of molecules detectable by cell-free biosensors by combining synthetic metabolic cascades with transcription factor-based networks. These hybrid cell-free biosensors are highly-sensitive and have a fast response and high-dynamic range. This work provides a foundation to engineer modular cell-free biosensors tailored for many applications.

synthetic biology↗