VIDEO - Membrane Proteins Workshop - From expression to purified protein: a P4-ATPase for CryoEM studies with LineMarie Christiansen
VIDEO - Membrane Proteins Workshop - From expression to purified protein: a P4-ATPase for CryoEM studies with LineMarie Christiansen
Speaker: LineMarie Christiansen, Aarhus University, Denmark
This lecture was part of the workshop, Structural Resolution of Membrane Proteins: From Expression to Sample , organised in May 2021 by the LINXS working group Membrane Proteins, which fall under the Integrative Structural Biology theme.
Abstract
The Saccharomyces cerevisiae P4-ATPase Neo1p is believed to function as a lipid flippase, translocating lipids towards the cytosolic leaflet of both Golgi and endosome membranes. In order to screen around both functional and structural studies to ascertain the exact role and function of Neo1p an efficient high-yield expression and purification protocol is vital. By overexpression in S. cerevisiae and purification based on a high-specificity tag we are left with a pure protein sample ideal for both structural and functional studies. For CryoEM detergent-exchange into LMNG can be easily performed, with some attention to detergent concentration