Lipid pumps and vesicular transport

If you are interested in working with lipid pumps and vesicular transport...Then join the Flippase Team!


The research groups of Associate Professors Rosa Lopez and Thomas Günther-Pomorski.
If you are interested in a project, please contact Rosa López (rlo@life.ku.dk) or Thomas Günther Pomorski (tgp@life.ku.dk).

A fundamental feature of eukaryotic cells is their compartmentalization into distinct organelles by lipid membranes. This allows different functions to occur efficiently and simultaneously in different parts of the cell. Among these functions are nutrient and ion transport, oxidative and photosynthetic phosphorylation, signal transduction, and electrical excitability. To fulfil these various functions, cells tightly regulate vesicular traffic between cellular compartments. Specific membrane pumps, termed lipid flippases, play an essential role in this process but how do they work and how are they regulated? To address these questions we perform an interdisciplinary research consisting of various techniques reaching from engineering, biochemistry, to modern cell biology and molecular analysis in yeast, plant and mammalian cells. Depending on your interest, several bachelor, master or PhD projects can be offered.

Below you find descriptions of some of the available projects.

Role of lipid pumps in vesicle biogenesis

Lipid flippases are involved in vesicle formation in the secretory pathway and interact with elements of the vesiculation machinery. The objective of this project is to analyse the interactions of two plant lipid flippases, localized to different compartments, with secretory pathway-related proteins. Applied techniques: RNA isolation, generation of a cDNA library, split-ubiquitin system setup, site-directed mutagenesis, transient expression of proteins in tobacco epidermal cells, bioimaging, membrane protein purification and co-immunoprecipitation.

Isolation and characterization of Arabidopsis flippases expressed in tobacco leaves

We have identified a plant lipid pump localised to vesicles when overexpressed in tobacco leaves. We would like to optimize a method to isolate these vesicles and biochemically characterize the flippase both directly in the vesicles and after purification and reconstitution in proteoliposomes. Applied techniques: cloning of genes using the USER system, site-directed mutagenesis, generation of 2A-linked fusion proteins, transient expression in tobacco epidermal leaf cells, bioimaging, membrane isolation, ATPase activity assay, phosphorylated intermediate studies, Immobilized Metal Affinity Chromatography (IMAC), lipid translocation assays.

Biochemical characterization of lipid pumps

We employ newly developed biophysical methods to measure the activity of lipid flippases. You will work on the purification and reconstitution of lipid flippases into liposomes of defined lipid composition. The lipid transport activity of the reconstituted proteins can be measured directly by using fluorescent lipid probes or by following the shape change in giant vesicles that can be observed with an optical microscope. 

Functional significance of lipid pumps in sperm

Many candidate lipid flippases are of medical relevance and their dysfunction contributes to the genesis of severe diseases in man. Here, you will work on the characterization of sperm lipid transporters and their physiological relevance. You will express these lipid pumps in yeast and mammalian cells and study their function in lipid and vesicular transport using fluorescence microscopy and flow cytometry.