Morris B J, Moffat B, Reid I A
Am J Physiol. 1979 Jun;236(6):E655-9. doi: 10.1152/ajpendo.1979.236.6.E655.
Dog angiotensinogen was purified 450-fold from the plasma of nephrectomized dogs by a simple four-step procedure involving precipitation between 1.5 and 2.3 M ammonium sulfate, gel filtration on Sephadex G-150, ion-exchange chromatography on DE-52 cellulose, and affinity chromatography on Concanavalin A-Sepharose. The purity of the final preparation was over 50%. The preparation of dog angiotensinogen had an apparent molecular weight of 80,000 determined by gel filtration on Sephadex G-100. Kinetic studies indicated that the Km of the reaction of dog renin with partially purified dog angiotensinogen (1,840 pmol/ml) was similar to that for the reaction with angiotensinogen in diluted dog plasma (1,820 pmol/ml). Thus the purification procedures employed did not alter the affinity of dog renin for the Leu10-Leu11 bond of dog angiotensinogen. Because the concentration of angiotensinogen in dog plasma is about 700 pmol/ml, a first order reaction with respect to substrate is indicated in vivo.
通过一个简单的四步程序，从肾切除犬的血浆中纯化犬血管紧张素原，纯化倍数达450倍。该程序包括在1.5至2.3M硫酸铵之间沉淀、在Sephadex G - 150上进行凝胶过滤、在DE - 52纤维素上进行离子交换色谱以及在伴刀豆球蛋白A - 琼脂糖上进行亲和色谱。最终制剂的纯度超过50%。通过在Sephadex G - 100上进行凝胶过滤测定，犬血管紧张素原制剂的表观分子量为80,000。动力学研究表明，犬肾素与部分纯化的犬血管紧张素原（1,840 pmol/ml）反应的Km与在稀释的犬血浆中与血管紧张素原反应的Km相似（1,820 pmol/ml）。因此，所采用的纯化程序并未改变犬肾素对犬血管紧张素原Leu10 - Leu11键的亲和力。由于犬血浆中血管紧张素原的浓度约为700 pmol/ml，表明体内底物反应为一级反应。
