Protein over-expression in Escherichia coli triggers adaptation analogous to antimicrobial resistance.
Protein over-expression in Escherichia coli triggers adaptation analogous to antimicrobial resistance.
复制标题
DOI：
10.1186/s12934-020-01462-6
复制
发表时间：
2021-01-11
影响因子：
6.4
通讯作者：
Doyle DA
中科院分区：
文献类型：
--
作者：
James J;Yarnall B;Koranteng A;Gibson J;Rahman T;Doyle DA
急诊coli pET系统是目前世界上应用最广泛的蛋白质过表达系统。它依赖于所有细胞产生靶蛋白的假设，并且通常认为，整合膜蛋白（IMP）过表达比其可溶性对应物更具毒性。使用GFP标记的蛋白质，可溶性或IMP靶标的高水平过表达导致> 99.9%的细胞损失，存活率仅<0.03%。选择压力产生三种表型：大绿色、大白色和小菌落变体。结果，在过夜培养物中，约50%的总细胞质量不产生蛋白质。表型的基因组测序揭示了导致T7 RNAP活性丧失或其转录下调的基因组突变。过表达过程是杀菌的，并且对于可溶性蛋白和膜蛋白都观察到。我们证明，这是外源蛋白在大肠杆菌中高水平过表达的行为。启动一系列事件，导致> 99.9%的细胞死亡。这些结果重新定义了我们对蛋白质过度生产的理解，并将其与抗生素耐药性发展中的适应性生存反应联系起来。
The E. coli pET system is the most widely used protein over-expression system worldwide. It relies on the assumption that all cells produce target protein and it is generally believed that integral membrane protein (IMP) over-expression is more toxic than their soluble counterparts. Using GFP-tagged proteins, high level over-expression of either soluble or IMP targets results in > 99.9% cell loss with survival rate of only < 0.03%. Selective pressure generates three phenotypes: large green, large white and small colony variants. As a result, in overnight cultures, ~ 50% of the overall cell mass produces no protein. Genome sequencing of the phenotypes revealed genomic mutations that causes either the loss of T7 RNAP activity or its transcriptional downregulation. The over-expression process is bactericidal and is observed for both soluble and membrane proteins. We demonstrate that it is the act of high-level over-expression of exogenous proteins in E. coli that sets in motion a chain of events leading to > 99.9% cell death. These results redefine our understanding of protein over-production and link it to the adaptive survival response seen in the development of antimicrobial resistance.
登录
查看更多内容
影响因子：
4.8
作者：
Liu, JY;Miller, PF;Olson, ER
通讯作者：
Olson, ER
影响因子：
3.5
作者：
Choi, Jae Young;Jang, Tae-Ho;Park, Hyun Ho
通讯作者：
Park, Hyun Ho
影响因子：
6.4
作者：
Li, Zhaopeng;Rinas, Ursula
通讯作者：
Rinas, Ursula
影响因子：
3.2
作者：
Edgar, R;Bibi, E
通讯作者：
Bibi, E
影响因子：
4.6
作者：
Kwon SK;Kim SK;Lee DH;Kim JF
通讯作者：
Kim JF