title: Inhibitory effect of Pb2+ on the transport cycle of the Na+,K+-ATPase.
abstract: The effect of Pb(2+) on the transport cycle of the Na(+),K(+)-ATPase was characterized in detail at a molecular level by combining electrical and biochemical measurements. Electrical measurements were performed by adsorbing purified membrane fragments containing Na(+),K(+)-ATPase on a solid-supported membrane. Upon adsorption, the Na(+),K(+)-ATPase was activated by carrying out concentration jumps of different activating substrates, for example, Na(+) and ATP. Charge movements following Na(+),K(+)-ATPase activation were measured in the presence of various Pb(2+) concentrations to investigate the effect of Pb(2+) on different ion translocating steps of the pump cycle. These charge measurements were then compared to biochemical measurements of ATPase activity in the presence of increasing Pb(2+) concentration. Our results indicate that Pb(2+) inhibits cycling of the enzyme, but it does not affect cytoplasmic Na(+) binding and release of Na(+) ions at the extracellular side at concentrations below 10 muM. To explain the inhibitory effect of Pb(2+) on the Na(+),K(+)-ATPase, we propose that Pb(2+) may interfere with the hydrolytic cleavage of the phosphorylated intermediate E(2)P, which occurs in the K(+)-related branch of the pump cycle.
authors.affiliation: Department of Evolutionary Biology Leo Pardi, University of Florence, 50125 Florence, Italy.
authors.forename: Elisa
authors.initials: E
authors.lastname: Gramigni
authors.forename: Francesco
authors.initials: F
authors.lastname: Tadini-Buoninsegni
authors.forename: Gianluca
authors.initials: G
authors.lastname: Bartolommei
authors.forename: Giacomo
authors.initials: G
authors.lastname: Santini
authors.forename: Guido
authors.initials: G
authors.lastname: Chelazzi
authors.forename: Maria Rosa
authors.initials: MR
authors.lastname: Moncelli
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D055664:Electrochemical Techniques / Q000379:methods; D004566:Electrodes; D006046:Gold / Q000737:chemistry; D017136:Ion Transport; D007854:Lead / Q000378:metabolism* / Q000633:toxicity; D010766:Phosphorylation; D011485:Protein Binding; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry / Q000378:metabolism*
chemical_list: D007854:Lead; D006046:Gold; D000255:Adenosine Triphosphate; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase