Click here to close Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly. We suggest using a current version of Chrome, FireFox, or Safari.
PLoS One January 1, 2011; 6 (7): e21901.
Show Gene links Show Anatomy links

Frog oocytes to unveil the structure and supramolecular organization of human transport proteins.

Bergeron MJ , Boggavarapu R , Meury M , Ucurum Z , Caron L , Isenring P , Hediger MA , Fotiadis D .

Structural analyses of heterologously expressed mammalian membrane proteins remain a great challenge given that microgram to milligram amounts of correctly folded and highly purified proteins are required. Here, we present a novel method for the expression and affinity purification of recombinant mammalian and in particular human transport proteins in Xenopus laevis frog oocytes. The method was validated for four human and one murine transporter. Negative stain transmission electron microscopy (TEM) and single particle analysis (SPA) of two of these transporters, i.e., the potassium-chloride cotransporter 4 (KCC4) and the aquaporin-1 (AQP1) water channel, revealed the expected quaternary structures within homogeneous preparations, and thus correct protein folding and assembly. This is the first time a cation-chloride cotransporter (SLC12) family member is isolated, and its shape, dimensions, low-resolution structure and oligomeric state determined by TEM, i.e., by a direct method. Finally, we were able to grow 2D crystals of human AQP1. The ability of AQP1 to crystallize was a strong indicator for the structural integrity of the purified recombinant protein. This approach will open the way for the structure determination of many human membrane transporters taking full advantage of the Xenopus laevis oocyte expression system that generally yields robust functional expression.

PubMed ID: 21760919
PMC ID: PMC3131388
Article link: PLoS One
Grant support: [+]

Species referenced: Xenopus laevis
Genes referenced: aqp1 myc slc15a1 slc1a1 slc5a1.2

Article Images: [+] show captions
References [+] :
Baur, A GABA(A) receptor of defined subunit composition and positioning: concatenation of five subunits. 2006, Pubmed, Xenbase