Photodynamic inactivation of the Na,K-ATPase occurs via different pathways.
The photodynamic, i.e., the light-induced, inactivation of the Na,K-ATPase in the presence of the sensitizer rose bengal was studied under different conditions. The shape of inactivation curves of the enzyme activity was analyzed as well as partial reactions of the pump cycle. Both experimental approaches showed the existence of two different time constants of inactivation of the ion pump, which reflect two pathways of a photodynamic modification. This is supported by the following observations: (1) The amplitude of the initial fast decay of enzyme activity was enhanced in the presence of D2O and reduced in the presence of the singlet oxygen scavenger imidazole. (Similar results were found for the SR Ca-ATPase.) (2) Contrary to the fast enzyme inactivation the slow process shows an inverse dose-rate behavior. (3) Inactivation of the partial reactions of Na+ -binding and of K+-binding to the membrane domain of the Na,K-ATPase showed only a single time constant, which corresponded to the slower time constant of enzyme inactivation. In the presence of high concentrations of singlet oxygen the fast time constant dominated the inactivation of the ATP-induced partial reaction for which the cytoplasmic domains of the enzyme play an important role. The data support the conclusion that fast inactivation is due to modification of the cytoplasmic domains and slow inactivation due to modifications of the membrane domain of the ion pumps.
Killig
F
F
Department of Biology, University of Konstanz, Konstanz, Germany.
Stark
G
G
Apell
H J
HJ
D000818:Animals; D017666:Deuterium Oxide / Q000737:chemistry; D004789:Enzyme Activation / Q000187:drug effects* / Q000528:radiation effects*; D005456:Fluorescent Dyes / Q000494:pharmacology*; D007700:Kinetics; D049449:Luminescence; D011188:Potassium / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects / Q000528:radiation effects; D017434:Protein Structure, Tertiary; D011817:Rabbits; D012395:Rose Bengal / Q000494:pharmacology*; D026082:Singlet Oxygen / Q000737:chemistry; D012964:Sodium / Q000737:chemistry / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry* / Q000378:metabolism
D005456:Fluorescent Dyes; D026082:Singlet Oxygen; D012395:Rose Bengal; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase; D017666:Deuterium Oxide; D011188:Potassium