Other Prokaryotic Systems

Halobacterium salinarum, a member of Archaebacteria, has a purple-colored plasma membrane that contains a complex of the bacterio-opsin protein (Bop) and its chromophore retinal in a 1:1 ratio.23 This complex, known as bacteriorhodopsin, forms highly ordered two-dimensional structures for which a high resolution structure was determined.24 Heterologous expression systems based on bacteriorhodopsin have been established for H. salinarum.25 Introduction of the human muscarinic M1 receptor and the human serotonin 5-HT2C receptor into this expression system disappointingly produced no signals with Western blotting.26 However, engineering of chimeric receptors between Bop and the yeast Ste2 receptor resulted in detectable expression levels.

Lactococcus lactis is a Gram-positive bacterium frequently applied to overexpression of integral membrane proteins.27 The most frequently used system is based on the nisin NisA promoter and the two regulatory trans-acting factors known as NisR and NisK.28 Most of the membrane proteins expressed to date are of pro-karyotic origin, typically ABC transporters and major facilitator subfamily (MSF) efflux pumps.27 Only a few eukaryotic targets have been expressed in L. lactis. The human Lys-Asp-Glu-Leu (KDEL) receptor with a 7TM topology has been expressed in a functional form although at moderate levels of less than 0.1% of the total membrane protein. In contrast, the yields for the 6TM yeast mitochondrial carrier proteins were in the range of 5%. No reports have been published for L. lactis-based GPCR expression but this approach may be worth investigating.

0 0

Post a comment