Oligomerization state of water channels and glycerol facilitators - Involvement of loop E

The major intrinsic protein (MIP) family includes water channels aquaporins (AQPs) and facilitators for small solutes such as glycerol (GlpFs), Veloci ty sedimentation on sucrose gradients demonstrates that heterologous AQPcic expressed in yeast or Xenopus oocytes behaves as an homotetramer when extr acted by n-octyl beta-D-glucopyranoside (OG) and as a monomer when extracte d by SDS, We performed an analysis of GlpF solubilized from membranes of Es cherichia coli or of mRNA-injected Xenopus oocytes, The GlpF protein extrac ted either by SDS or by nondenaturing detergents, OG and Triton X-100, exhi bits sedimentation coefficients only compatible with a monomeric form of th e protein in micelles, We then substituted in loop E of AQPcic two amino ac ids predicted to play a role in the functional/structural properties of the MIPs, In two expression systems, yeast and oocytes, the mutant AQPcic-S205 D is monomeric in OG and in SDS, The A209K mutation does not modify the tet rameric form of the heterologous protein in OG, This study shows that the s erine residue at position 205 is essential for AQPcic tetramerization, Beca use the serine in this position is highly conserved among aquaporins and sy stematically replaced by an acid aspartic in GlpFs, we postulate that glyce rol facilitators are monomers whereas aquaporins are organized in tetramers , Our data suggest that the role of loop E in MIP properties partly occurs through its ability to allow oligomerization of the proteins.