MEASURING EQUILIBRIUM CONSTANTS FOR THE DISSOCIATION OF LIGANDS FROM P-TYPE ATPases: A new strategy to determine conformers and measure equilibrium constants in P-type ATPases using a photoactivatable phospholipidic probe.

IRENE MANGIALAVORI, MARIELA FERREIRA GOMES, MARÍA F. PIGNATARO, EMANUEL E STREHLER AND JUAN PABLO F.C

Buzios- Rio de Janeiro. Brasil

Workshop; Workshop del Latin American Postgraduate Program of Biophysics Course; 2009

Latin American Postgraduate Program of Biophysics

  Information about the structure and assembly of the transmembrane domain of an integral membrane protein can be obtained from the analysis of the lipid–protein interactions. We developed a hydrophobic photolabeling method to study the non-covalent interactions between PMCA and surrounding phospholipids under different experimental conditions that lead to known conformations. Equilibrium constants for the dissociation of ligands from PMCA complexes are measured through the changing of transmembrane conformations. The method is sensitive, precise and requires a very low amount of protein allowing to its application to membrane proteins that cycle through different conformations like P-type ATPases