CONICET | Buscador de Institutos y Recursos Humanos

In this work, we characterize the stability against the detergent sodium dodecyl sulfate (SDS) of the ATP binding domain (ATPBD) of the Cu(I) transport ATPase from Archaeoglobus fulgidus CopA, a hiperthermophilic α-helical membrane protein that is involved in transporting Cu+ throughout biological membranes. To accomplish this, we carry out experiments registering the intrinsic fluorescence of the unique tryptophan resildue as a function of the detergent concentration. A decrease in the fluorescence was observed in the range of 0.03 ? 1.6 mM of SDS with 3 different phases in the downfall. This decrease was proving to be reversible by dialysis and even by dilution of the sample, which recover the original fluorescence of the protein. We also observed that intrinsic anisotropy increased in the range 0.2 - 1 mM SDS, indicating that the SDS-denatured protein acquire a rigid structure. Finally, we explore the interaction of CopA-ATPBD with the fluorescent probe 1-anilino-naphthalene-8-sulfonate (ANS), observing an initial increase of the ANS fluorescence until 0.3 mM SDS followed by decrease up to a constant value at 2-3 mM SDS. We can hypothesize that SDS-induced unfolding of CopA-ATPBD proceeds through an intermediate state that acquire an expanded -molten globule like- globular structure, holding the ANS in a more hydrophobic environment.