Charla Invitada: Detergent-membrane interaction. Is it useful to investigate lipid domains?

P.M. RODI; A.M. GENNARO

Salta

Congreso; XXXIX Annual Meeting of the Argentinean Biophysical Society and 3rd Latin American Protein Society Meeting; 2010

Latin American Protein Society- SAB

In the last years we have been researching about detergent solubility of the erythrocyte membrane (1,2). The initial goal was to analyze the properties of detergent resistant domains (DRMs), but besides of studying the insoluble material obtained at one specific detergent concentration, we became interested in analyzing what happened when the samples were treated with increasing detergent concentrations. We performed detailed quantifications of phosphorus and cholesterol content in the insoluble material obtained from intact and cholesterol depleted human and bovine erythrocytes, treated with different Triton X-100 and CHAPS concentrations, at 4º or 37ºC. The raft hypothesis proposes that membranes are not homogeneous, but have ordered domains (rafts) which act as platforms for protein coalescence. Rafts are suggested to be in a liquid ordered state and to be insoluble in cold detergent solutions. The identity of rafts and DRM is under discussion. Usual descriptions of membrane solubilization (3) would predict a steady decrease in the amount of insoluble lipids as a function of detergent concentration for a homogeneous liquid disordered membrane. However, if the membrane were not uniform but presented macroscopic ordered domains, the presence of some kind of plateaus would be expected in the solubility plots. Our results show in all cases a monotonous decrease of insoluble lipids as detergent concentration is increased, and no plateaus could be detected. Thus, our results do not support the hypothesis of the presence of macroscopic ordered domains in the erythrocyte membrane. However, if the size of rafts were small enough, their existence would be compatible with our solubility plots. We have also demonstrated the existence of strong interactions between the DRMs and the erythrocyte citoskeleton. This find is an important issue not only in the DRM isolation protocol but also reflects a significant interaction between citoskeleton and putative lipid domains.