Toxin studies using an integrated biophysical and structural biology approach.

Clostridial neurotoxins, such as botulinum and tetanus, are generally thought to invade neural cells through a process of high affinity binding mediated by gangliosides, internalization via endosome formation, and subsequent membrane penetration of the catalytic domain activated by a pH drop in the endosome. This surface recognition and internalization process is still not well understood with regard to what specific membrane features the toxins target, the intermolecular interactions between bound toxins, and the molecular conformational changes that occur as a result of pH lowering. In an effort to elucidate the mechanism of tetanus toxin binding and permeation through the membrane a simple yet representative model was developed that consisted of the ganglioside G{sub tlb} incorporated in a bilayer of cholesterol and DPPC (dipalmitoylphosphatidyl choline). The bilayers were stable over time yet sensitive towards the binding and activity of whole toxin. A liposome leakage study at constant pH as well as with a pH gradient, to mimic the processes of the endosome, was used to elucidate the effect of pH on the toxin's membrane binding and permeation capability. Topographic imaging of the membrane surface, via in situ tapping mode AFM, provided nanoscale characterization of the toxin's binding location and pore formation activity

PenQuest Volume 1, Number 2

Table of Contents for this Volume: Untitled by Julie Ambrose Night by Judith Gallo Untitled by Judy Gozdur the shamans by Charles Riddles Untitled by Jerry Connell Untitled by Laura Woods Untitled by LEMA Wicked Bird by Laura Jo Last Untitled by Rick Dentos Untitled by Jeni Moody Untitled by Bettie W. Kwibs Untitled by Joann Stagg The Protector Stood by Laura Jo Last Visions of Salome by Charles Riddles Untitled by Thomas Tutten Kennesaw Line by Don Ova-Dunaway Stone Blood by Mary Ellen C. Wofford Untitled by Roger Whitt Jr. Untitled by C. Wingate Untitled by Doug Dorey Untitled by Karen Blumberg Untitled by Beverly Oviatt Untitled by Virginia Shrader The Crapulous Credo of Charles C. by Charles Riddles the brave and the true by David Reed Untitled by Charles Gutierrez Canoe Creek by Patricia Kraft Untitled by Linda Bobinger The Man in the Iron Lung by Patricia Kraft Untitled by Roger Whitt, Jr. Childish Things by Kathleen Gay Untitled by Joseph Avanzini The Lover by Mary S. Aken Untitled by Ann Harrington And He Taketh Away by David Reed Untitled by Mary Graham Untitled by Melody A. Cummons Untitled by Karen Blumberg To The Poets by Judith Gallo Untitled by Ann Harringto

Lipid membranes on nanostructured silicon.

A unique composite nanoscale architecture that combines the self-organization and molecular dynamics of lipid membranes with a corrugated nanotextured silicon wafer was prepared and characterized with fluorescence microscopy and scanning probe microscopy. The goal of this project was to understand how such structures can be assembled for supported membrane research and how the interfacial interactions between the solid substrate and the soft, self-assembled material create unique physical and mechanical behavior through the confinement of phases in the membrane. The nanometer scale structure of the silicon wafer was produced through interference lithography followed by anisotropic wet etching. For the present study, a line pattern with 100 nm line widths, 200 nm depth and a pitch of 360 nm pitch was fabricated. Lipid membranes were successfully adsorbed on the structured silicon surface via membrane fusion techniques. The surface topology of the bilayer-Si structure was imaged using in situ tapping mode atomic force microscopy (AFM). The membrane was observed to drape over the silicon structure producing an undulated topology with amplitude of 40 nm that matched the 360 nm pitch of the silicon structure. Fluorescence recovery after photobleaching (FRAP) experiments found that on the microscale those same structures exhibit anisotropic lipid mobility that was coincident with the silicon substructure. The results showed that while the lipid membrane maintains much of its self-assembled structure in the composite architecture, the silicon substructure indeed influences the dynamics of the molecular motion within the membrane