Samis J A, Elce J S
Department of Biochemistry, Queen's University, Kingston, Ontario, Canada.
Thromb Haemost. 1989 Apr 25;61(2):250-3.
With a view to understanding the function of calpain I (EC 3.4.22.17) in vivo, the localization of the enzyme was studied by immunoelectron microscopy in human erythrocytes. Thin sections of the cells embedded in Spurr's resin were exposed to solutions of monoclonal anti-calpain I or control antibodies, biotin antimouse IgG antibodies, and streptavidin-gold. Most of the calpain I (93%) was found to be distributed throughout the cytoplasm, and only 7% of the gold particles were associated with the erythrocyte membrane. Erythrocytes were Ca2+-loaded by means of the calcium ionophore A23187, and the rise in intracellular [Ca2+] was demonstrated both by crenation of the cells, and by activation of calpain which was detected by immunoblotting. The proportions of cytosolic and membrane-bound gold labelling were, however, not altered by Ca2+-loading. These results are not consistent with the hypothesis that activation of calpain requires membrane-binding.
为了了解钙蛋白酶I（EC 3.4.22.17）在体内的功能，通过免疫电子显微镜研究了该酶在人红细胞中的定位。将包埋在Spurr树脂中的细胞薄片暴露于单克隆抗钙蛋白酶I抗体或对照抗体、生物素抗小鼠IgG抗体和链霉亲和素-金溶液中。发现大部分钙蛋白酶I（93%）分布在整个细胞质中，只有7%的金颗粒与红细胞膜相关。通过钙离子载体A23187使红细胞加载Ca2+，细胞皱缩和免疫印迹检测到的钙蛋白酶激活均证明细胞内[Ca2+]升高。然而，Ca2+加载并未改变胞质和膜结合金标记的比例。这些结果与钙蛋白酶激活需要膜结合的假设不一致。
