Influence of environmental factors on molecular transport through bacterial membranes

Abstract

Bacterial membranes act as protective barriers and help to regulate molecular interactions between a cell and its surrounding environment. External chemical and physical influences have the potential to alter the properties of bacterial membranes and therefore impact the viability of the cell. This can stem from natural or seasonal changes to the local environment (e.g., temperature, pH, and salinity), or even deliberate application of an antimicrobial agent. Regardless, understanding exactly how such external stimuli influence bacterial membrane properties is of fundamental importance, both in terms of basic microbiology as well as for designing pharmaceutical interventions. Experimentally, this is a non-trivial task as this requires selective isolation of a signal arising from the membrane, which is typically buried in the overwhelming background response of the surrounding bulk environment. In particular, our lab has previously developed the surface-sensitive nonlinear optical method, second harmonic light scattering (SHS), as a means of interrogating molecular interactions at the membrane surfaces of living cells, even for multimembrane systems (e.g., Gram-negative bacteria). In this dissertation, time-resolved SHS was employed to study a variety of membrane properties across two separate projects, including 1) chemical and physical induced changes in membrane permeability and 2) temperature-induced membrane permeability changes. Specifically, in the first project (Chapter 4), the influence of the signaling molecule, indole, on the permeability of the bacterial cytoplasmic membrane was quantified. It was revealed that the interaction of indole with the tryptophan specific transporting protein, Mtr permease, resulted in enhanced passive diffusion across the membrane. For the second project (Chapter 5), we examined the influence of temperature on the rate of passive diffusion across a membrane, both in model systems (liposomes) and in living cells (E. coli). For both bacterial and liposome systems, increasing temperatures resulted in a modest increase in passive diffusion rates across the membrane. However, when the temperature range included a phase transition, passive diffusion increased by an order of magnitude. Therefore, by monitoring transport rate in relation to temperature, membrane phase transitions can be quantitatively determined based on the characteristic discontinuities in the measured trend.