Department of Molecular Biosciences, University of Texas at Austin, Austin, TX, USA.
John Ring LaMontagne Center for Infectious Diseases, The University of Texas at Austin, Austin, TX, USA.
Mol Microbiol. 2021 Jul;116(1):16-28. doi: 10.1111/mmi.14700. Epub 2021 Feb 25.
Protein folding is central to both biological function and recombinant protein production. In bacterial expression systems, which are easy to use and offer high protein yields, production of the protein of interest in its native fold can be hampered by the limitations of endogenous posttranslational modification systems. Disulfide bond formation, entailing the covalent linkage of proximal cysteine amino acids, is a fundamental posttranslational modification reaction that often underpins protein stability, especially in extracytoplasmic environments. When these bonds are not formed correctly, the yield and activity of the resultant protein are dramatically decreased. Although the mechanism of oxidative protein folding is well understood, unwanted or incorrect disulfide bond formation often presents a stumbling block for the expression of cysteine-containing proteins in bacteria. It is therefore important to consider the biochemistry of prokaryotic disulfide bond formation systems in the context of protein production, in order to take advantage of the full potential of such pathways in biotechnology applications. Here, we provide a critical overview of the use of bacterial oxidative folding in protein production so far, and propose a practical decision-making workflow for exploiting disulfide bond formation for the expression of any given protein of interest.
蛋白质折叠是生物功能和重组蛋白生产的核心。在细菌表达系统中，由于其易于使用且能够产生高蛋白质产量，因此在其天然折叠状态下生产目标蛋白可能会受到内源性翻译后修饰系统的限制。二硫键的形成涉及到邻近半胱氨酸氨基酸的共价连接，是一种基本的翻译后修饰反应，通常可以提高蛋白质的稳定性，尤其是在细胞外环境中。当这些键不能正确形成时，所得蛋白质的产量和活性会大大降低。尽管氧化蛋白折叠的机制已经得到很好的理解，但在细菌中表达含有半胱氨酸的蛋白质时，不想要的或不正确的二硫键形成常常是一个障碍。因此，在进行蛋白质生产时，考虑到原核生物中二硫键形成系统的生物化学特性非常重要，以便在生物技术应用中充分利用这些途径的潜力。在这里，我们对迄今为止细菌氧化折叠在蛋白质生产中的应用进行了批判性的综述，并提出了一种实用的决策工作流程，用于利用二硫键形成来表达任何给定的目标蛋白。
