Flik G, Verbost P M, Atsma W, Lucu C
Department of Animal Physiology, Faculty of Science, University of Nijmegen, The Netherlands.
J Exp Biol. 1994 Oct;195:109-22. doi: 10.1242/jeb.195.1.109.
A procedure was developed for the preparation of inside-out vesicles from plasma membranes isolated from the branchial epithelium of the green shore crab Carcinus maenas (L.). Procedures normally applied to fish branchial epithelium required the introduction of an additional hypotonic shock to obtain a preparation containing 22% inside-out vesicles, 33% right-side-out vesicles and 45% leaky membrane fragments. In such membrane preparations, the first direct evidence for uphill (against a [Ca2+] gradient) ATP-dependent and Na(+)-gradient-dependent Ca2+ transport in crustacean gills was found. The affinity for Ca2+ of the ATP-driven Ca2+ transporter was 149 nmol l-1 and that of the Na+/Ca2+ exchanger was 1.78 mumol l-1; the Vmax values were 1.73 and 9.88 nmol min-1 mg-1 protein respectively. The relative importance of these carriers for Ca2+ transport in the branchial epithelium of the crab is evaluated on the basis of their calcium kinetics.
已开发出一种从绿岸蟹（Carcinus maenas (L.)）鳃上皮分离的质膜制备内翻囊泡的方法。通常应用于鱼鳃上皮的方法需要额外引入低渗休克，以获得一种包含22%内翻囊泡、33%外翻囊泡和45%渗漏膜片段的制剂。在这种膜制剂中，首次发现了甲壳类动物鳃中存在依赖ATP（逆[Ca2+]梯度）和依赖Na+梯度的Ca2+上坡转运的直接证据。ATP驱动的Ca2+转运体对Ca2+的亲和力为149 nmol l-1，Na+/Ca2+交换体对Ca2+的亲和力为1.78 μmol l-1；Vmax值分别为1.73和9.88 nmol min-1 mg-1蛋白质。根据这些载体的钙动力学评估了它们在蟹鳃上皮中Ca2+转运的相对重要性。
