Chauvelot-Moachon L, Tallet F, Durlach-Misteli C, Giroud J P
Department of Pharmacology, Hôpital Cochin, Paris, France.
J Pharmacol Methods. 1988 Aug;20(1):15-28. doi: 10.1016/0160-5402(88)90012-5.
Propranolol binding to human alpha 1-acid glycoprotein (AAG) delipidated by two methods is described. Commercial AAG (99% pure) was either precipitated by ethanol-acetone and then washed by ether, or it was precipitated by ethanol. Binding capacity was quantified by the product n x Ka where n denotes the number of binding sites and Ka the association constant (M-1). Propranolol binding to nondelipidated AAG (n x Ka = 0.113 +/- 0.013 microM-1) was clearly increased after precipitation by ethanol-acetone (n x Ka = 0.386 +/- 0.109 microM-1) or precipitation by ethanol (n x Ka = 0.312 +/- 0.096 microM-1). Binding capacity potentiation cannot be due to modification of AAG microheterogeneity forms, as two-dimensional gel electrophoresis pattern of AAG in presence of concanavalin A was not altered after both methods. Recombination of precipitated AAGs with supernatant dry residue resulted in the abrogation of observed potentiation. Moreover, addition of a polar lipid, linoleic acid, (from 30 to 300 microM) strongly inhibited propranolol binding. These results indicated that glycoprotein precipitation by ethanol provided a simple method to further study binding inhibitors associated with isolated AAG.
描述了通过两种方法脱脂的普萘洛尔与人α1-酸性糖蛋白（AAG）的结合情况。市售AAG（99%纯）要么用乙醇-丙酮沉淀，然后用乙醚洗涤，要么用乙醇沉淀。结合能力通过n×Ka产物进行定量，其中n表示结合位点的数量，Ka表示缔合常数（M-1）。普萘洛尔与未脱脂AAG的结合（n×Ka = 0.113±0.013 μM-1）在经乙醇-丙酮沉淀（n×Ka = 0.386±0.109 μM-1）或乙醇沉淀（n×Ka = 0.312±0.096 μM-1）后明显增加。结合能力的增强并非由于AAG微不均一性形式的改变，因为两种方法处理后，伴刀豆球蛋白A存在下AAG的二维凝胶电泳图谱未发生变化。沉淀的AAG与上清液干残渣重新组合导致观察到的增强作用消失。此外，添加极性脂质亚油酸（30至300 μM）强烈抑制普萘洛尔的结合。这些结果表明，用乙醇沉淀糖蛋白提供了一种简单的方法来进一步研究与分离的AAG相关的结合抑制剂。
