Generation of a vector suite for protein solubility screening
Generation of a vector suite for protein solubility screening
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DOI：
10.3389/fmicb.2014.00067
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发表时间：
2014-02-25
影响因子：
5.2
通讯作者：
Oppezzo, Pablo
中科院分区：
文献类型：
--
作者：
Correa, Agustin;Ortega, Claudia;Oppezzo, Pablo
重组蛋白表达已成为学术和生物技术项目的宝贵工具。利用可溶性蛋白生产的高通量筛选技术，以可溶性和均质的状态生产了无数的目标蛋白，从而能够实现进一步的研究。数百种条件的评估需要使用高通量克隆和筛选方法。在这里，我们描述了一个新的多功能载体套件，致力于表达的重组蛋白（RP）的溶解性问题的改善。该载体套件允许将相同的PCR产物平行克隆到12种不同的表达载体中，以评估在不同启动子强度、不同融合标签以及不同溶解度增强子蛋白下的蛋白质表达。此外，我们建议使用一种新的融合蛋白，这似乎是一个有用的溶解度增强剂。最重要的是，我们在这项工作中提出了一个经济和有用的载体套件，以快速跟踪不同RP的溶解度。我们还提出了一种新的溶解度增强蛋白，可以包括在经典的表达条件下不溶的RP的表达的评估。
Recombinant protein expression has become an invaluable tool for academic and biotechnological projects. With the use of high-throughput screening technologies for soluble protein production, uncountable target proteins have been produced in a soluble and homogeneous state enabling the realization of further studies. Evaluation of hundreds conditions requires the use of high-throughput cloning and screening methods. Here we describe a new versatile vector suite dedicated to the expression improvement of recombinant proteins (RP) with solubility problems. This vector suite allows the parallel cloning of the same PCR product into the 12 different expression vectors evaluating protein expression under different promoter strength, different fusion tags as well as different solubility enhancer proteins. Additionally, we propose the use of a new fusion protein which appears to be a useful solubility enhancer. Above all we propose in this work an economic and useful vector suite to fast track the solubility of different RP We also propose a new solubility enhancer protein that can be included in the evaluation of the expression of RP that are insoluble in classical expression conditions.