The natural polyphenol, epigallocatechin-3-gallate, inhibits the Plasma Membrane Ca2+-ATPase by favoring the phosphoenzyme conformation

MANGIALAVORI, IRENE CECILIA; FERRIERA GOMES, MARIELA; RINALDI, DÉBORA; REY, OSVALDO; SAFFIOTI, NICOLÁS ANDRÉS; ROSSI JUAN PABLO; ROSSI ROLANDO

Tucumán

Congreso; III Latin American Federation of Biophysical Societies (LAFeBS) ? IX IberoAmerican Congress of Biophysics ? XLV Reunion Anual SAB 2016; 2016

Latin American Federation of Biophysical Societies (LAFeBS)

Epigallocatechin 3-gallate (EGCG) is the major polyphenol componentof green tea. This compound is believed to be the main responsible formany of the health benefits associated with green tea. EGCG benefi-cial effects include antioxidant, anti-inflammatory and neuroprotectiveeffects. EGCG produces alterations in intracellular Ca2+ homeostasiswhich could be linked to the function of the Sarcoplasmic ReticulumCalcium Pump (SERCA). The maintenance of intracellular calciumlevels is fine-tuned by the Plasma Membrane Ca2+-ATPase (PMCA).Therefore, we investigated the effect of different flavan-3-ols and wentinto detail about the EGCG effect on PMCA.We performed measurements of the Ca2+-ATPase activity on PMCApurified from human red blood cells and of Ca2+ flux on HEK293Tcells that overexpress PMCA4. We evaluated PMCA ATPase activityin the presence of catechin (C), epicatechin (EC) and epigallocatechin.C and EC showed no effect up to 100 µM. However, EGCG showed astrong inhibition with a K0.5 of 0.032 ± 0.003 µM. Under similar conditions,EGCg showed an increase of the phosphorylated intermediatewhich was found to be ADP sensitive, suggesting that EGCG could stabilizethe E1P conformation on the reaction cycle of hydrolysis of ATPby PMCA.We performed docking assays of EGCG as ligand and PMCA structuremodels as the receptor. PMCA models were obtained by homologymodelling on SERCA crystallographic structures in different conformations.Our results showed that ECGC binds mainly to the closedconformations of the enzyme. Particularly the E1P conformation formsthe lowest free energy biding complexes with the ligand establishinghydrogen bonds with both the P and N domains of the pump.On the other hand, Ca2+ eflux in HEK293 cells is inhibited by EGCGin vivo showing its physiological relevance on PMCA activity.With grants of CONICET, ANPCYT and UBACYT.