VIDEO - Membrane Proteins Workshop - An yeast GFP-based platform for functional and structures investigation of SLC transporters with David Drew
VIDEO - Membrane Proteins Workshop - An yeast GFP-based platform for functional and structures investigation of SLC transporters with David Drew
Speaker: David Drew, Stockholm University
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
Recombinant expression screening of eukaryotic membrane proteins for functional and structural investigation is still a trial-and-error process. We have found that cloning gene-strings by homologuous recombination into a 2u vector S. cerevisiae expression vector and then detecting membrane protein expression and stability by working with GFP-fusions, enables many constructs to be rapidly tested. We have found an excellent correlation between the best-behaving membrane proteins in yeast to those produced by transient transfection in mammalian HEK293 cells. Here, I will outline these GFP-based methods with a focus on ion and sugar SLC transporters.