TY - BOOK

T1 - Analysis of Protein-Membrane Interactions

T2 - A Liposomal Approach

AU - Kemmer, Gerdi Christine

PY - 2013

Y1 - 2013

N2 - Cellular membranes are complex structures, consisting of hundreds of different lipids and proteins. These membranes act as barriers between distinct environments, constituting hot spots for many essential functions of the cell, including signaling, energy conversion, and transport. These functions are implemented by soluble proteins reversibly binding to, as well as by integral membrane proteins embedded in, cellular membranes. The activity and interaction of these proteins is furthermore modulated by the lipids of the membrane. Here, liposomes were used as model membrane systems to investigate interactions between proteins and lipids. First, interactions of soluble proteins with membranes and specific lipids were studied, using two proteins: Annexin V and Tma1. The protein was first subjected to a lipid/protein overlay assay to identify candidate interaction partners in a fast and efficient way. Discovered interactions were then probed on the level of the membrane using liposome-based assays. In the second part, a transmembrane protein was investigated. Assays to probe activity of the plasma membrane ATPase (Arabidopsis thaliana H+ -ATPase isoform 2 (AHA2)) in single liposomes using both giant vesicles and immobilized proteoliposomes were established. To enable investigation of proton pumping on the single-liposome level, two lipid-linked pH-sensors were synthesized, purified, and characterized

AB - Cellular membranes are complex structures, consisting of hundreds of different lipids and proteins. These membranes act as barriers between distinct environments, constituting hot spots for many essential functions of the cell, including signaling, energy conversion, and transport. These functions are implemented by soluble proteins reversibly binding to, as well as by integral membrane proteins embedded in, cellular membranes. The activity and interaction of these proteins is furthermore modulated by the lipids of the membrane. Here, liposomes were used as model membrane systems to investigate interactions between proteins and lipids. First, interactions of soluble proteins with membranes and specific lipids were studied, using two proteins: Annexin V and Tma1. The protein was first subjected to a lipid/protein overlay assay to identify candidate interaction partners in a fast and efficient way. Discovered interactions were then probed on the level of the membrane using liposome-based assays. In the second part, a transmembrane protein was investigated. Assays to probe activity of the plasma membrane ATPase (Arabidopsis thaliana H+ -ATPase isoform 2 (AHA2)) in single liposomes using both giant vesicles and immobilized proteoliposomes were established. To enable investigation of proton pumping on the single-liposome level, two lipid-linked pH-sensors were synthesized, purified, and characterized

UR - https://soeg.kb.dk/permalink/45KBDK_KGL/fbp0ps/alma99122699925205763

M3 - Ph.D. thesis

BT - Analysis of Protein-Membrane Interactions

PB - Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen

ER -