Journal
PROTEIN SCIENCE
Volume 14, Issue 12, Pages 3048-3056Publisher
WILEY
DOI: 10.1110/ps.051689905
Keywords
eukaryotic membrane proteins; mitochondrial carriers; nisin-inducible expression system; Lactococcus lactis; transport
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Funding
- Medical Research Council [MC_U105663139] Funding Source: Medline
- MRC [MC_U105663139] Funding Source: UKRI
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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 I I 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.
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