Kracke G R, O'Neal S G, Chacko G K
J Membr Biol. 1981;63(1-2):147-56. doi: 10.1007/BF01969455.
The (Na+ + K+)-ATPase of garfish olfactory nerve axon plasma membrane was purified about sixfold by treatment of the membrane with sodium dodecyl sulfate followed by sucrose density gradient centrifugation. The estimated molecular weights of the two major polypeptide components of the enzyme preparation on sodium dodecyl sulfate gels were 110,000 and 42,000 daltons, which were different from those of the corresponding peptides of rabbit kidney (Na+ + K+)-ATPase. No carbohydrate was detected in the 42,000-dalton component either by the periodic acid-Schiff reagent or by the more sensitive concanavalin A-peroxidase staining procedure. The molecular properties of the garfish (Na+ + K+)-ATPase, such as the Km for ATP, pH optimum, energies of activation, Na and K ion dependence and vanadium inhibition, were, however, similar to those of the kidney enzyme. The partially purified garfish (Na+ + K+)-ATPase was reconstituted into phospholipid vesicles by a freeze-thaw-sonication procedure. The reconstituted enzyme was found to catalyze a time and ATP dependent 22Na+ transport. The ratio of 22Na+ pumped to ATP hydrolyzed was about 1; under the same reconstitution and assay conditions, eel electroplax (Na+ + K+)-ATPase, however, gave a 22Na+ pumped to ATP hydrolyzed ratio of nearly 3.
用十二烷基硫酸钠处理雀鳝嗅神经轴突质膜，然后进行蔗糖密度梯度离心，可将其（Na⁺ + K⁺）-ATP酶纯化约6倍。在十二烷基硫酸钠凝胶上，该酶制剂的两种主要多肽成分的估计分子量分别为110,000和42,000道尔顿，这与兔肾（Na⁺ + K⁺）-ATP酶相应肽段的分子量不同。用高碘酸-席夫试剂或更灵敏的伴刀豆球蛋白A-过氧化物酶染色法，均未在42,000道尔顿的成分中检测到碳水化合物。然而，雀鳝（Na⁺ + K⁺）-ATP酶的分子特性，如对ATP的米氏常数、最适pH值、活化能、对Na和K离子的依赖性以及钒抑制作用，与肾酶相似。通过冻融-超声处理程序，将部分纯化的雀鳝（Na⁺ + K⁺）-ATP酶重组成磷脂囊泡。发现重组酶可催化与时间和ATP相关的²²Na⁺转运。泵出的²²Na⁺与水解的ATP的比率约为1；然而，在相同的重组和测定条件下，鳗鱼电板（Na⁺ + K⁺）-ATP酶泵出的²²Na⁺与水解的ATP的比率接近3。
