tailieunhanh - Báo cáo khoa học: The molecular chaperone a-crystallin incorporated into red cell ghosts protects membrane Na/K-ATPase against glycation and oxidative stress

a-Crystallin, a molecular chaperone and lens structural protein protects soluble enzymes against heat-induced aggregation and inactivation by a variety of molecules. In this study we investigated the chaperone function ofa-crys-tallininamorephysiological systeminwhicha-crystallinwas incorporated into red cell ghosts . Its ability to protect the intrinsic membrane protein Na/K-ATPase from external stresses was studied. Red cell ghosts were created by lysing the red cells and removing cytoplasmic contents by size-exclusion chromatography. . | Eur. J. Biochem. 270 2605-2611 2003 FEBS 2003 doi The molecular chaperone a-crystallin incorporated into red cell ghosts protects membrane Na K-ATPase against glycation and oxidative stress Barry K. Derham1 J. Clive Ellory2 Anthony J. Bron1 and John J. Harding1 1Nuffield Laboratory of Ophthalmology University of Oxford UK laboratory of Physiology University of Oxford UK a-Crystallin a molecular chaperone and lens structural protein protects soluble enzymes against heat-induced aggregation and inactivation by a variety of molecules. In this study we investigated the chaperone function of a-crys-tallin in a more physiological system in which a-crystallin was incorporated into red cell ghosts . Its ability to protect the intrinsic membrane protein Na K-ATPase from external stresses was studied. Red cell ghosts were created by lysing the red cells and removing cytoplasmic contents by sizeexclusion chromatography. The resulting ghost cells retain Na K-ATPase activity. a-Crystallin was incorporated in the cells on resealing and the activity of Na K-ATPase assessed by ouabain-sensitive 86Rb uptake. Incubation with fructose hydrogen peroxide and methylglyoxal compounds that have been implicated in diabetes and cataract formation were used to test inactivation of the Na Kpump. Intracellular a-crystallin protected against the decrease in ouabain sensitive 86Rb uptake and therefore against inactivation induced by all external modifiers in a dose-dependent manner. Keywords a-crystallin ghost cells glycation Na K-ATPase oxidation. Na K-ATPase is a highly conserved ubiquitous membrane protein. The enzyme is composed of three subunits the alpha subunit w 113 kDa binds ATP and sodium and potassium ions and contains the phosphorylation site. The smaller beta subunit w 35 kDa glycoprotein is necessary for activity of the complex and the gamma subunit w 10 kDa is involved with modulation of Na K-ATPase. Several isoforms of both alpha and beta subunits