谷氨酸棒杆菌（乳酸发酵短杆菌）ftsZ基因在大肠杆菌中的过表达。

Overexpression of the ftsZ gene from Corynebacterium glutamicum (Brevibacterium lactofermentum) in Escherichia coli.

作者信息

Honrubia-Marcos María Pilar, Ramos Angelina, Gil José A

机构信息

Area de Microbiología, Departamento de Ecología, Genética y Microbiología, Facultad de Biología, Universidad de León, Spain.

出版信息

Can J Microbiol. 2005 Jan;51(1):85-9. doi: 10.1139/w04-105.

DOI:10.1139/w04-105

PMID:15782238

Abstract

Our goal in this work was to overexpress the essential cell division FtsZ protein from Corynebacterium glutamicum (Brevibacterium lactofermentum) (FtsZCG) in Escherichia coli to produce anti-FtsZCG polyclonal antibodies. Previous results from our laboratory showed that ftsZCG was not expressed in E. coli in a sufficient amount to purify FtsZCG. However, when ftsZCG (without upstream sequences) was transcriptionally fused to the T7 promoter, different truncated FtsZCG proteins (28-32 kDa) were overexpressed in E. coli, and in all cases, stop codons were created because of DNA deletions or rearrangements. Nevertheless, we were able to overexpress and purify an N-terminally hexa-His-tagged FtsZCG from uninduced E. coli cells carrying a pET-28a(+) derivative, yielding about 5 mg of 98% pure protein per 100-mL culture.

摘要

我们这项工作的目标是在大肠杆菌中过表达谷氨酸棒杆菌（乳酸发酵短杆菌）的必需细胞分裂蛋白FtsZ（FtsZCG），以产生抗FtsZCG多克隆抗体。我们实验室之前的结果表明，ftsZCG在大肠杆菌中表达量不足，无法纯化FtsZCG。然而，当ftsZCG（无上游序列）与T7启动子进行转录融合时，不同的截短FtsZCG蛋白（28 - 32 kDa）在大肠杆菌中过表达，并且在所有情况下，由于DNA缺失或重排产生了终止密码子。尽管如此，我们能够从携带pET - 28a(+)衍生物的未诱导大肠杆菌细胞中过表达并纯化N端带有六聚组氨酸标签的FtsZCG，每100 mL培养物可产生约5 mg纯度为98%的蛋白质。