Preparation and Properties of Vesicles with Highly Controlled Lipid Asymmetry

Suny At Stony Brook, Stony Brook NY

Investigators

Abstract

This award by the Biomaterials program in the Division of Materials Research to State University of New York at Stony Brook is to investigate basic structure and function of membranes using asymmetric lipid vesicles. Lipid asymmetry, a difference in the lipid composition in the inner and outer layers of a biological membrane, is a prominent and crucial property of the lipid bilayer of many cell membranes. Artificial vesicles (liposomes) composed of lipid bilayers are biomaterials that have proven invaluable models of biological membranes. Using cyclodextrins, preliminary studies by the investigator indicated the feasibility of preparing asymmetric lipid vesicles by selectively introducing lipids into the outer monolayer of a lipid vesicle via lipid exchange. This project will systematically study the conditions that control the extent of lipid exchange using a wide range of lipids, and will investigate important basic questions about membranes. Conditions that provide most efficient lipid binding by cyclodextrins will be identified, and then conditions will be optimized for maximal lipid exchange while maintaining asymmetry. The second goal is to study biologically-important physical properties of vesicles that control asymmetry and its impact on membrane structure. The first property to be studied is coupling between the physical state of lipids in the outer and inner leaflets as a function of lipid structure. This interleaflet coupling may transduce signals from the outside to the inside of membranes. These experiments are expected to distinguish between alternate hypotheses for how lipids and proteins promote or inhibit interleaflet coupling. The second property to be studied will be lipid transverse diffusion (lipid flip-flop) between leaflets, and how this is impacted by lipid structure and membrane proteins. It is important, both to define the conditions and identify proteins that can be used while maintaining stable lipid asymmetry, and to understand what membrane protein sequences affect transverse diffusion. This project will have a broad impact on career development of future scientists, including minority students, at the educational level by training both graduate and undergraduate students. They will receive specialized training in a wide variety of biochemical techniques used to study membrane proteins and lipid, including spectroscopic techniques that emphasize the fluorescence and fluorescence quenching approaches. The students will also be trained in proper conduct of scientific studies, writing and speaking, and preparing them for careers in field of biological/biophysical research and/or teaching. This project should allow facile applications of the methods by other labs, with applications that have a significant impact upon human (and animal) health, including drug delivery applications. Nanomaterial applications in which molecular-width layers with different chemical compositions are needed will be developed by this research project. Living cells are surrounded by membranes composed of membrane lipids (specialized fat molecules) and proteins. These membranes control the uptake of nutrients into cells, and the communication between cells and their environment. Cell membranes are very complex, consisting of thousands of types of different proteins and lipids. Artificial membrane vesicles, composed of small variety of membrane lipids and proteins which form an envelope similar to a cell membrane, are important tools for understanding how membranes function. Natural membranes have two layers of membrane-lipids and are asymmetric, with different types of lipids in each layer, but a key limitation in the utility of artificial membranes has been their lack of lipid asymmetry. The recently developed method to prepare asymmetric artificial membranes by the investigator will be studied in detail to allow it to be used with a wide variety of different lipids and proteins. The use artificial membranes will greatly expand the knowledge base on membranes. This project will also have a broad impact on career development of future scientists, including minority students. At the educational level, both graduate and undergraduate students (including via contacts with other local institutions) will be trained in the conduct of research, experimental principles, and the science of studying membrane structure and function including spectroscopic techniques. These students will also be trained in proper conduct of scientific studies, scientific writing and speaking, and preparing them for careers in field of scientific research and teaching.