Insights into the mechanism of erythrocyte Na+/K+-ATPase inhibition by nitric oxide and peroxynitrite anion.
Evidence shows that Na(+)/K(+)-ATPase from kidney, brain and liver is inhibited by nitric oxide (NO) and peroxynitrite anion (ONO(2) (-)), but the mechanism is unknown. The aim of the present work was to study the inhibitory effect of NO and ONO(2) (-) on erythrocyte Na(+)/K(+)-ATPase. Erythrocyte membranes were isolated from male Wistar rats by hypotonic washing. The membranes (free from haemoglobin) were incubated for Na(+)/K(+)-ATPase activity measurement at various concentrations of ATP in the presence or absence of 400 microM SNAP (an NO donor) or 100 microM SIN-1 (an ONO(2)(-) donor). At these concentrations, SNAP and SIN-1 released about the same amount (100 microM) of NO or ONO(2)(-), respectively, as monitored by measuring NO(2)(-) + NO(3)(-). Both SNAP and SIN-1 decreased V(max) by ca. 75% but they did not decrease the apparent affinity of the Na(+)/K(+)-ATPase for the substrate (a decrease of K(m) was even observed after SNAP treatment). The pattern of this inhibition is compatible either with oxidation of SH groups directly involved in ATP binding but in a way that is not surmountable by increasing the substrate concentration ("non-competitive") or with oxidation of SH groups located outside the active site of the enzyme but important for the activity of the enzyme.
Muriel
Pablo
P
Sección Externa de Farmacología, CINVESTAV-I.P.N., Apdo. Postal 14-740, México 07000, D. F. México. pamuriel@mail.cinevestav.mx
Castañeda
Gilberto
G
Ortega
Mónica
M
Noël
François
F
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000187:drug effects* / Q000201:enzymology; D066298:In Vitro Techniques; D008297:Male; D008981:Molsidomine / Q000031:analogs & derivatives* / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D010396:Penicillamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D030421:Peroxynitrous Acid / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism*
D004791:Enzyme Inhibitors; D020030:Nitric Oxide Donors; C110881:S-nitro-N-acetylpenicillamine; D030421:Peroxynitrous Acid; D009569:Nitric Oxide; C002385:linsidomine; D000255:Adenosine Triphosphate; D008981:Molsidomine; D000254:Sodium-Potassium-Exchanging ATPase; D010396:Penicillamine