Highly efficient export of a disulfide-bonded protein to the periplasm and medium by the Tat pathway using CyDisCo in Escherichia coli

IF 3.9 3区 生物学 Q2 MICROBIOLOGY MicrobiologyOpen Pub Date : 2023-03-08 DOI:10.1002/mbo3.1350
Klaudia Arauzo-Aguilera, Mirva J. Saaranen, Colin Robinson, Lloyd W. Ruddock
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Abstract

High-value heterologous proteins produced in Escherichia coli that contain disulfide bonds are almost invariably targeted to the periplasm via the Sec pathway as it, among other advantages, enables disulfide bond formation and simplifies downstream processing. However, the Sec system cannot transport complex or rapidly folding proteins, as it only transports proteins in an unfolded state. The Tat system also transports proteins to the periplasm, and it has significant potential as an alternative means of recombinant protein production because it transports fully folded proteins. Most of the studies related to Tat secretion have used the well-studied TorA signal peptide that is Tat-specific, but this signal peptide also tends to induce degradation of the protein of interest, resulting in lower yields. This makes it difficult to use Tat in the industry. In this study, we show that a model disulfide bond-containing protein, YebF, can be exported to the periplasm and media at a very high level by the Tat pathway in a manner almost completely dependent on cytoplasmic disulfide formation, by other two putative Tat SPs: those of MdoD and AmiC. In contrast, the TorA SP exports YebF at a low level.

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在大肠杆菌中使用CyDisCo通过Tat途径高效地将二硫结合蛋白输出到外质和培养基中
大肠杆菌中产生的含有二硫键的高价值外源蛋白几乎总是通过Sec途径靶向周质,因为它具有其他优点,可以形成二硫键并简化下游加工。然而,Sec系统不能运输复杂或快速折叠的蛋白质,因为它只能运输未折叠状态的蛋白质。Tat系统也将蛋白质运输到外周质,由于它运输完全折叠的蛋白质,因此它作为重组蛋白质生产的替代手段具有重要的潜力。大多数与Tat分泌相关的研究都使用了经过充分研究的Tat特异性TorA信号肽,但这种信号肽也容易诱导目标蛋白的降解,导致产量降低。这使得在行业中很难使用Tat。在这项研究中,我们发现一种含二硫键的模型蛋白YebF可以通过Tat途径以非常高的水平输出到外周质和培养基中,其方式几乎完全依赖于细胞质二硫的形成,通过另外两个假定的Tat SPs: MdoD和AmiC。相比之下,TorA SP在较低水平上输出YebF。
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来源期刊
MicrobiologyOpen
MicrobiologyOpen MICROBIOLOGY-
CiteScore
8.00
自引率
0.00%
发文量
78
审稿时长
20 weeks
期刊介绍: MicrobiologyOpen is a peer reviewed, fully open access, broad-scope, and interdisciplinary journal delivering rapid decisions and fast publication of microbial science, a field which is undergoing a profound and exciting evolution in this post-genomic era. The journal aims to serve the research community by providing a vehicle for authors wishing to publish quality research in both fundamental and applied microbiology. Our goal is to publish articles that stimulate discussion and debate, as well as add to our knowledge base and further the understanding of microbial interactions and microbial processes. MicrobiologyOpen gives prompt and equal consideration to articles reporting theoretical, experimental, applied, and descriptive work in all aspects of bacteriology, virology, mycology and protistology, including, but not limited to: - agriculture - antimicrobial resistance - astrobiology - biochemistry - biotechnology - cell and molecular biology - clinical microbiology - computational, systems, and synthetic microbiology - environmental science - evolutionary biology, ecology, and systematics - food science and technology - genetics and genomics - geobiology and earth science - host-microbe interactions - infectious diseases - natural products discovery - pharmaceutical and medicinal chemistry - physiology - plant pathology - veterinary microbiology We will consider submissions across unicellular and cell-cluster organisms: prokaryotes (bacteria, archaea) and eukaryotes (fungi, protists, microalgae, lichens), as well as viruses and prions infecting or interacting with microorganisms, plants and animals, including genetic, biochemical, biophysical, bioinformatic and structural analyses. The journal features Original Articles (including full Research articles, Method articles, and Short Communications), Commentaries, Reviews, and Editorials. Original papers must report well-conducted research with conclusions supported by the data presented in the article. We also support confirmatory research and aim to work with authors to meet reviewer expectations. MicrobiologyOpen publishes articles submitted directly to the journal and those referred from other Wiley journals.
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