Partial purification and characterization of the (Ca2+ + Mg2+)-ATPase from squid optic nerve plasma membrane.
A membrane fraction enriched in axolemma was obtained from optic nerves of the squid (Sepiotheutis sepioidea) by differential centrifugation and density gradient fractionation. The preparation showed an oligomycin- and NaN3-insensitive (Ca2+ + Mg2+)-ATPase activity. The dependence of the ATPase activity on calcium concentration revealed the presence of two saturable components. One had a high affinity for calcium (K1 1/2 = 0.12 microM) and the second had a comparatively low affinity (K2 1/2 = 49.5 microM). Only the high-affinity component was specifically inhibited by vanadate (K1 = 35 microM). Calmodulin (12.5 micrograms/ml) stimulated the (Ca2+ + Mg2+)-ATPase by approx. 50%, and this stimulation was abolished by trifluoperazine (10 microM). Further treatment of the membrane fraction with 1% Nonidet P-40 resulted in a partial purification of the ATPase about 15-fold compared to the initial homogenate. This (Ca2+ + Mg2+)-ATPase from squid optic nerve displays some properties similar to those of the uncoupled Ca2+-pump described in internally dialyzed squid axons, suggesting that it could be its enzymatic basis.
D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase; D000252:Calcium-Transporting ATPases / Q000302:isolation & purification* / Q000378:metabolism; D002147:Calmodulin / Q000494:pharmacology; D049832:Decapodiformes; D003902:Detergents; D009441:Neurilemma / Q000201:enzymology*; D009900:Optic Nerve / Q000201:enzymology*; D014639:Vanadium / Q000494:pharmacology
D002147:Calmodulin; D003902:Detergents; D014639:Vanadium; D017301:Ca(2+) Mg(2+)-ATPase; D000252:Calcium-Transporting ATPases