Open position : Linking proteins and their binding partners to lipid nanodiscs

Linking proteins and their binding partners to lipid nanodiscs
Joint project to be executed by an Experienced Researcher (ER/postdoc) between Intana, SiChem, and the EMBL.
Many interactions between cellular biomolecules take place at the cell membrane, but in vitro assays, mimicking membrane bound interactions have been complicated to establish.
In a novel biochemical model, we propose to develop a method based on linkers for attaching proteins and their binding partners to lipid nanodiscs (Intana, 8 months). On one hand, Click chemistry will be employed to produce linkers featuring azido or tetrazine moieties (SiChem, 8 months). To increase residence time and decrease motility in the nanodisc, photo-crosslinkable lipids such as diazirines and their covalent coupling to other lipids in the disk might be used. Proteins of interest including those important in sphingosine metabolism will be equipped with an artificial amino acid providing a strained cyclooctyne or cyclooctene group (Plass et al., 2011, 2012). Proteins will be expressed in E. coliand purified at EMBL (EMBL, 2 months). Alternatively, SNAP -, CLIP-, or HALO-Tags will be used to attach proteins to nanodiscs featuring the respective benzylguanine etc lipid linkers. Proof-of-concept will demonstrate the use of the method to investigate protein-drug interactions.

Figure legend: 2-Dimensional Interaction Trap.
Proteins and interaction partners are covalently attached to membranes via enzymatic fusion to lipids anchoring in the nanodisc (Snap- and Clip- Tag, left side) or via Click chemistry. Interaction will be monitored by FRET and drugs interfering with this interaction (and FRET) will be tested by simple addition.