肠系膜明串珠菌B-512FM葡糖转移酶的简易纯化方法。

A facile purification of Leuconostoc mesenteroides B-512FM dextransucrase.

作者信息

Fu D T, Robyt J F

机构信息

Department of Biochemistry and Biophysics, Iowa State University, Ames 50011.

出版信息

Prep Biochem. 1990;20(2):93-106. doi: 10.1080/00327489008050182.

DOI:10.1080/00327489008050182

PMID:2146604

Abstract

Leuconostoc mesenteroides NRRL B-512F has been mutated by treatment with N-nitrosoguanidine. The resulting mutant (designated as B-512FM) produces 300 times as much enzyme as the parent strain. B-512FM dextransucrase was treated extensively with Sigma crude dextranase, followed by column chromatography on Bio-Gel A-5m. The purified dextransucrase had a specific activity of 84 IU/mg, a 100-fold purification with 42% yield, and was shown by SDS-PAGE to have a single protein of molecular weight of 158,000 with dextransucrase activity. The procedure has been used to produce purified enzyme for sequencing. The molecular weight of 158,000 agrees with that calculated from its amino acid sequence.

摘要

肠膜明串珠菌NRRL B - 512F经N - 亚硝基胍处理后发生了突变。产生的突变体（命名为B - 512FM）产生的酶量是亲本菌株的300倍。用Sigma粗葡聚糖酶对B - 512FM葡聚糖蔗糖酶进行了广泛处理，随后在Bio - Gel A - 5m上进行柱色谱分离。纯化后的葡聚糖蔗糖酶比活性为84 IU/mg，纯化倍数为100倍，产率为42%，SDS - PAGE显示其具有一条分子量为158,000且具有葡聚糖蔗糖酶活性的单一蛋白质条带。该方法已用于生产用于测序的纯化酶。158,000的分子量与根据其氨基酸序列计算出的分子量一致。