Functional expression of eukaryotic membrane proteins in Lactococcus lactis

The overproduction of eukaryotic membrane proteins is a major impediment in their structural and functional characterization. Here we have used the nisin‐inducible expression system of Lactococcus lactis for the overproduction of 11 mitochondrial transport proteins from yeast. They were expressed at high levels in a functional state in the cytoplasmic membrane. The results also show that the level of expression is influenced by the N‐terminal regions of the transporters. Expression levels were improved >10‐fold either by replacing or truncating these regions or by adding lactococcal signal peptides. The observed expression levels are now compatible with a realistic exploration of crystallization conditions. The lactococcal expression system may be used for the high‐throughput functional characterization of eukaryotic membrane proteins and structural genomics.