Biology subjects

Samuel, I. D. W.

Publications and source records attributed to Samuel, I. D. W..

3 recordsLinked to original sources

bioRxiv2026

Understanding the role of membrane lipids in mechanism of antimicrobial photodynamic therapy in Escherichia coli

Antimicrobial photodynamic therapy (aPDT) is a promising alternative to antibiotics, yet the molecular factors determining bacterial susceptibility remain unclear. This study investigates the critical role of bacterial lipids, particularly cardiolipin (CL) in the aPDT response of Escherichia coli using methylene blue as a photosensitizer. Through genetic deletion ({Delta}clsABC) and chemical modification via mannitol supplementation, we demonstrate that reduced CL levels significantly enhance bacterial sensitivity, leading to an additional reduction in viability exceeding 3 log10. Quantitative lipidomics (MS,GC) confirmed substantial CL depletion and altered fatty acid saturation. Interestingly, while live CL-deficient cells were more vulnerable, biomimetic giant unilamellar vesicles (GUVs) with higher CL content showed greater susceptibility to photo-oxidation. These findings suggest that CL-rich microdomains in living bacteria act as functional scaffolds for stress-defense systems rather than mere targets for oxidative damage. Modulating membrane lipid composition thus represents a novel strategy to potentiate aPDT efficacy.

microbiology↗

bioRxiv2024

Analysis and control of untemplated DNA polymerase activity for guided synthesis of kilobase-scale DNA sequences

DNA polymerases are complex molecular machines able to replicate genetic material using a template-driven process. While the copying function of these enzymes is well established, their ability to perform untemplated DNA synthesis is less well characterized. Here, we explore the ability of DNA polymerases to synthesize DNA fragments in the absence of template. We use long-read nanopore sequencing and real-time PCR to observe the synthesis of pools of DNA products derived from a diverse set of natural and engineered DNA polymerases across varying temperatures and buffer compositions. We detail the features of the DNA fragments generated, enrichment of select sequence motifs, and demonstrate that the sequence composition of the synthesized DNA may be altered by modifying environmental conditions. This work provides an extensive data set to better discern the process of untemplated DNA polymerase activity and may support its potential repurposing as a technology for the guided synthesis of DNA sequences on the kilobase-scale and beyond.

synthetic biology↗

bioRxiv2019

Unveiling the multi-step solubilization mechanism of single vesicles by detergents.

The solubilization of membranes by detergents is critical for many technological applications and has become widely used in biochemistry research to induce cell rupture, extract cell constituents, and to purify, reconstitute and crystallize membrane proteins. The thermodynamic details of solubilization have been extensively investigated, but the kinetic aspects remain poorly understood. Here we used a combination of single-vesicle Forster resonance energy transfer (svFRET), fluorescence correlation spectroscopy and quartz-crystal microbalance with dissipation monitoring to access the real-time kinetics and elementary solubilization steps of sub-micron sized vesicles, which are inaccessible by conventional diffraction-limited optical methods. Real-time injection of a non-ionic detergent, Triton X, induced biphasic solubilization kinetics of surface-immobilized vesicles labelled with the Dil/DiD FRET pair. The nanoscale sensitivity accessible by svFRET allowed us to unambiguously assign each kinetic step to distortions of the vesicle structure comprising an initial fast vesicle-swelling event followed by slow lipid loss and micellization. We expect the svFRET platform to be applicable beyond the sub-micron sizes studied here and become a unique tool to unravel the complex kinetics of detergent-lipid interactions.

biophysics↗