Obstructed lateral diffusion of lipids and membrane proteins in polymer -tethered bilayers

Miranda A Deverall, Purdue University

Abstract

The exploration of hindered diffusion is crucial to understanding membrane organization and dynamics. Obstacles embedded in the lipid bilayer hinder the diffusion of lipids and proteins. Such obstacles are created by binding membrane proteins to the cytoskeleton or extracellular matrix, by small lipid domains, or by crowding of mobile membrane proteins. The main experimental problem is that the area fraction of obstacles in cellular membranes can only be roughly estimated. This research overcomes this limitation by exploring three different aspects of membrane dynamics related to obstructed diffusion in model membrane systems: obstacle-induced obstructed diffusion of lipids and proteins by small obstacles of tethered lipids, obstructed diffusion of lipids in the outer (obstacle-free leaflet of the bilayer), and lateral mobility of tethered lipids in crowded conditions. A polymer-supported bilayer is used for these studies with the tethered lipids acting as small obstacles. It is observed that increasing tethering concentrations cause decreasing lateral mobility of both lipids and transmembrane proteins, and that these lipid-anchored molecules act as immobile point obstacles. Comparing tracking experiments on lipid and bacteriorhodopsin (BR) tracer molecules reveal that diffusion is dependent upon tracer size. The lateral mobility of lipids in the outer leaflet (obstacle-free) were measured and surprisingly had identical diffusion coefficients to lipids in the inner monolayer (obstacle-containing). This study shows for the first time the qualitative and quantitative degree of obstructed diffusion caused by tethered lipids and that non-membrane spanning obstacles, if bound to polymeric material, create a virtual obstacle in the opposite leaflet of the bilayer. It also provides insight into the nature of the cytoskeleton-membrane attachment and membrane dynamics. Crowded diffusion studies on lipopolymers were conducted by varying the amount of lipopolymer in the outer leaflet of a polymer-tethered bilayer. This revealed that as lipopolymer concentration in the outer leaflet increases, the lateral mobility of the lipopolymer decreases demonstrating that the lipopolymers behave like mobile obstacles and that the interaction of the hydrophilic moieties of the lipopolymers is extremely important to diffusion processes.

Degree

Ph.D.

Advisors

Naumann, Purdue University.

Subject Area

Biochemistry

COinS