Effects of volatile anesthetic on the Ca2+-ATPase activation by dimerization. Distance-dependent quenching analysis and fluorescence energy transfer studies.
The phenomenological distance-dependent quenching (DDQ) model was employed to investigate the character of the interaction between volatile anesthetics (VAs) and the plasma membrane Ca2+-ATPase (PMCA). The simultaneous analysis of the frequency-domain and steady-state data of tryptophan (Trp) fluorescence quenching by a VA points to a specific character of the apparent quenching effect of the VA, possibly arising from a significant contribution of static quenching. The apparent contributions of both static and dynamic quenching may be due to VA binding in the PMCA, which results in the modification of the conformational substates of the enzyme. To characterize further the molecular consequences of VA binding, we investigated its effects on the process of PMCA activation by self-association. VA shifted the equilibrium from enzyme dimers to monomers, as monitored by the loss of fluorescence energy transfer. The shift was apparently due to the VA-induced decrease in the affinity of PMCA molecules for self-association. Addition of a large molecular mass dextran to increase the proximity between enzyme monomers induced re-association of the VA-impaired PMCA, while the Ca2+-ATPase activity was not recovered. The results are congruent with a dual VA effect on PMCA, a shift in the monomer/dimer equilibrium, and an inactivation of both monomers and dimers.
Lopez
M M
MM
Department of Anesthesiology/Critical Care Medicine, The Johns Hopkins School of Medicine, Baltimore, Maryland, USA. MXL53@PSU.EDU
Zelent
B
B
Kosk-Kosicka
D
D
D000777:Anesthetics / Q000737:chemistry / Q000494:pharmacology*; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D019281:Dimerization; D004789:Enzyme Activation / Q000187:drug effects; D004910:Erythrocyte Membrane / Q000201:enzymology; D005470:Fluorometry; D006801:Humans; D007530:Isoflurane / Q000494:pharmacology*; D008956:Models, Chemical; D014364:Tryptophan / Q000737:chemistry
D000777:Anesthetics; D014364:Tryptophan; D007530:Isoflurane; D000252:Calcium-Transporting ATPases