by Keyword: Transmembrane proteins

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Sanmartí-Espinal, M., Galve, R., Iavicoli, P., Persuy, M. A., Pajot-Augy, E., Marco, M. P., Samitier, J., (2016). Immunochemical strategy for quantification of G-coupled olfactory receptor proteins on natural nanovesicles Colloids and Surfaces B: Biointerfaces 139, 269-276

Cell membrane proteins are involved in a variety of biochemical pathways and therefore constitute important targets for therapy and development of new drugs. Bioanalytical platforms and binding assays using these membrane protein receptors for drug screening or diagnostic require the construction of well-characterized liposome and lipid bilayer arrays that act as support to prevent protein denaturation during biochip processing. Quantification of the protein receptors in the lipid membrane arrays is a key issue in order to produce reproducible and well-characterized chips. Herein, we report a novel immunochemical analytical approach for the quantification of membrane proteins (i.e., G-protein-coupled receptor, GPCR) in nanovesicles (NVs). The procedure allows direct determination of tagged receptors (i.e., c-myc tag) without any previous protein purification or extraction steps. The immunochemical method is based on a microplate ELISA format and quantifies this tag on proteins embedded in NVs with detectability in the picomolar range, using protein bioconjugates as reference standards. The applicability of the method is demonstrated through the quantification of the c-myc-olfactory receptor (OR, c-myc-OR1740) in the cell membrane NVs. The reported method opens the possibility to develop well-characterized drug-screening platforms based on G-coupled proteins embedded on membranes.

Keywords: Bioelectronic nose, Competitive ELISA, G-protein-coupled receptors quantification, Natural vesicles, Olfactory receptors, Transmembrane proteins