通过精氨酸-琼脂糖亲和色谱法从大豆中纯化结节特异性尿酸酶。

Purification of nodule-specific uricase from soybean by arginine-sepharose affinity chromatography.

作者信息

Larsen K

机构信息

Department of Molecular Biology and Plant Physiology, University of Aarhus, Denmark.

出版信息

Prep Biochem. 1990;20(1):1-9. doi: 10.1080/00327489008050173.

DOI:10.1080/00327489008050173

PMID:2367346

Abstract

A method to purify uricase from soybean root nodules is described. The separation uses a single affinity chromatography step on Arginine-Sepharose, which was constructed by coupling L-Arginine to Activated CH-Sepharose 4B. Crude extracts were loaded onto small columns of Arginine-Sepharose and a significant retardation of uricase was observed. With a re-run of the fraction containing maximal uricase activity on the same column highly purified enzyme was obtained. Analysis by SDS-polyacrylamide gel electrophoresis revealed two protein bands. Analytical isoelectric focusing showed two isoforms of uricase, one dominating with pI 9.0 and a minor with pI 7.0.

摘要

本文描述了一种从大豆根瘤中纯化尿酸酶的方法。该分离方法采用在精氨酸-琼脂糖凝胶上进行单步亲和色谱，精氨酸-琼脂糖凝胶是通过将L-精氨酸偶联到活化的CH-琼脂糖凝胶4B上构建而成。将粗提物加载到精氨酸-琼脂糖凝胶的小柱上，观察到尿酸酶有明显的滞留现象。将含有最大尿酸酶活性的馏分在同一柱上重新运行，可获得高度纯化的酶。SDS-聚丙烯酰胺凝胶电泳分析显示有两条蛋白带。分析性等电聚焦显示尿酸酶有两种同工型，一种以pI 9.0为主，另一种以pI 7.0为辅。