title: Addition of subunit γ, K⁺ ions, and lipid restores the thermal stability of solubilized Na,K-ATPase.
abstract: Differential scanning calorimetry (DSC) was applied to ascertain the effect caused by K⁺, Na⁺, ATP, detergent, DPPC, DPPE, and subunit γ on the thermostability of Na,K-ATPase. The enthalpy variation (ΔH) for the thermal denaturation of the membrane-bound is twice the ΔH value obtained for solubilized Na,K-ATPase. Denaturation occurs in five steps for membrane-bound against three steps for the solubilized enzyme, therefore a multi-step unfolding process. In the presence of Na⁺, the melting temperature is 61.6°C, and the ΔH is lower as compared with the ΔH obtained in the presence or in the absence of K⁺. Addition of ATP does not alter the transition temperatures significantly, but the shape of the curve is modified. Subunit γ probably stabilizes Na,K-ATPase in the beginning of thermal unfolding, and different amounts of detergents in the solubilized sample change the protein stability. Reconstitution of Na,K-ATPase into a liposome shows that lipids exert a protector effect. These results reveal differences on the thermostability depending on the conformation of Na,K-ATPase. They are relevant because it allows a comparison with future studies, e.g. how the composition of the membrane interferes on the stability of Na, K-ATPase, elucidating the importance of the lipid type contained in cell membrane.
authors: Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, FFCLRP-USP, Depto. Química, 14040-901 Ribeirão Preto, SP, Brazil.
authors: Juliana Sakamoto
authors: JS
authors: Yoneda
authors: Carolina Fortes
authors: CF
authors: Rigos
authors: Pietro
authors: P
authors: Ciancaglini
mesh_terms: D000818:Animals; D002152:Calorimetry, Differential Scanning; D002414:Cations, Monovalent / Q000737:chemistry / Q000378:metabolism; D002462:Cell Membrane / Q000201:enzymology*; D011188:Potassium / Q000737:chemistry* / Q000378:metabolism; D011489:Protein Denaturation; D017510:Protein Folding; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D018892:Proton-Motive Force; D011817:Rabbits; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry* / Q000378:metabolism; D012995:Solubility
chemical_list: D002414:Cations, Monovalent; D021122:Protein Subunits; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium