猪肾碱性磷酸酶。纯化方法及分子特性
Alkaline phosphatase from pig kidney. Method of purification and molecular properties.
作者信息
Wachsmuth E D, Hiwada K
出版信息
Biochem J. 1974 Jul;141(1):273-82. doi: 10.1042/bj1410273.
Abstract
Alkaline phosphatase (EC 3.1.3.1) from pig kidney brush-border membranes was solubilized from membrane precipitates by butan-1-ol at a critical pH of 7.0. The 12000-fold purification procedure included (NH(4))(2)SO(4) precipitation, DEAE-and TEAE-cellulose chromatography, Sephadex G-200 gel filtration and neuraminidase digestion followed by DEAE-cellulose chromatography. The purified protein contained 20% (w/w) carbohydrate and had mol.wt. 150000-156000 as estimated by Sephadex filtration and ultracentrifuge analysis. It was a tetrameric glycoprotein consisting of identical subunits, and it had a molecular activity at 25 degrees C of 2600s(-1) per tetramer. Its concentration in kidney was estimated to be 8.5-8.8mg/kg.
摘要
猪肾刷状缘膜中的碱性磷酸酶（EC 3.1.3.1）在临界pH值7.0下，通过丁醇从膜沉淀物中溶解出来。12000倍的纯化过程包括硫酸铵沉淀、DEAE-和TEAE-纤维素色谱、Sephadex G-200凝胶过滤以及神经氨酸酶消化，随后进行DEAE-纤维素色谱。纯化后的蛋白质含有20%（w/w）的碳水化合物，通过Sephadex过滤和超速离心分析估计其分子量为150000 - 156000。它是一种由相同亚基组成的四聚体糖蛋白，在25℃时每四聚体的分子活性为2600 s⁻¹。其在肾脏中的浓度估计为8.5 - 8.8 mg/kg。
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