Conformational Changes Produced by ATP Binding to the Plasma Membrane Calcium Pump

IRENE MANGIALAVORI; MARIELA FERREIRA GOMES; NICOLAS ANDRÉS SAFFIOTI; RODOLFO GONZALES LEBRERO; ROLANDO ROSSI; JUAN PABLO ROSSI

JOURNAL OF BIOLOGICAL CHEMISTRY

AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

Año: 2013 vol. 288 p. 31030 - 31041

0021-9258

The effects of lipids on membrane proteins are likely to be complex and unique for each membrane protein. Here we studied different detergent/phosphatidylcholine reconstitution media and tested their effects on Plasma Membrane Ca2+-pump (PMCA). We found that Ca2+-ATPase activity shows a biphasic behaviour with respect to the detergent/phosphatidylcholine ratio. Moreover, the maximal Ca2+-ATPase activity largely depends on the length and the unsaturation degree of the hydrocarbon chain. Using Static Light Scattering and Fluorescence Correlation Spectroscopy, we monitored the changes in hydrodynamic radius of detergent/phosphatidylcholine particles during the micelle-vesicle transition. We found that, when PMCA is reconstituted in mixed micelles, neutral phospholipids increase the enzyme turnover. The biophysical changes associated to the transition from mixed micelles to bicelles, increase the time of residence of the phosphorylated intermediate (EP) decreasing the enzyme turnover. Molecular Dynamics simulations analysis of the interactions between PMCA and the phospholipid bilayer in which it is embedded show that in the 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) bilayer, charged residues of the protein are trapped in the hydrophobic core. Conversely, in the 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) bilayer, the overall hydrophobic?hydrophilic requirements of the protein surface are fulfilled the best, reducing the thermodynamic cost of exposing charged residues to the hydrophobic core. The apparent mismatch produced by a DOPC thicker bilayer, could be a structural foundation to explain its functional implication on PMCA.