School of Life Sciences, Nantong University, Nantong, 226019, China.
School of Life Sciences, Nantong University, Nantong, 226019, China.
Protein Expr Purif. 2021 Sep;185:105896. doi: 10.1016/j.pep.2021.105896. Epub 2021 May 1.
DNase is a powerful tool for a series of molecular biology applications. Developing a strategy for large-scale production of DNase with high purity and activity is critical for scientific research. In this study, a previously uncharacterized gene with nuclease activity was found in Trichogramma pretiosum genome. Pichia pastoris GS115 was preferred as the host to overcome the issues related to prokaryotic expression. Under the optimal conditions, the activity of T. pretiosum DNase (Tp-DNase) reached 1940 U/mL of culture supernatant in fed-batch fermentation. Using ion-exchange chromatography and adsorption chromatography, Tp-DNase was produced with a purity of >99% and molecular weight of 45 kDa. In vitro DNA degradation experiments showed that Tp-DNase could effectively degrade dsDNA, and its activity was slightly higher than that of bovine pancreas DNase I under the same conditions. Moreover, Tp-DNase can be used to eliminate nucleic acid contamination and improve the accuracy of nucleic acid detection.
DNase 是一系列分子生物学应用的有力工具。开发一种大规模生产高纯度和高活性的 DNase 的策略对于科学研究至关重要。在这项研究中，在 Trichogramma pretiosum 基因组中发现了一种具有核酸酶活性的先前未表征的基因。毕赤酵母 GS115 被选为宿主，以克服与原核表达相关的问题。在最佳条件下，在分批补料发酵中，T. pretiosum DNase（Tp-DNase）的活性达到了 1940 U/mL 的培养上清液。通过离子交换层析和吸附层析，Tp-DNase 的纯度达到了>99%，分子量为 45 kDa。体外 DNA 降解实验表明，Tp-DNase 可以有效地降解 dsDNA，其活性在相同条件下略高于牛胰腺 DNase I。此外，Tp-DNase 可用于消除核酸污染，提高核酸检测的准确性。
