Recently, the study of chitinases has become an important target of numerous research projects due to their potential for applications, such as biocontrol pest agents. Plant chitinases from carnivorous plants of the genus Drosera are most aggressive against a wide range of phytopathogens. However, low solubility or insolubility of the target protein hampered application of chitinases as biofungicides. To obtain plant chitinase from carnivorous plants of the genus Drosera in soluble form in E.coli expression strains, three different approaches including dialysis, rapid dilution, and refolding on Ni-NTA agarose to renaturation were tested. The developed « Rapid dilution » protocol with renaturation buffer supplemented by 10% glycerol and 2M arginine in combination with the redox pair of reduced/oxidized glutathione, increased the yield of active soluble protein to 9.5 mg per 1 g of wet biomass. A structure-based removal of free cysteines in the core domain based on homology modeling of the structure was carried out in order to improve the soluble of chitinase. One improved chitinase variant (C191A/C231S/C286T) was identified which shows improved expression and solubility in E. coli expression systems compared to wild type. Computational analyzes of the wild-type and the improved variant revealed overall higher fluctuations of the structure while maintaining a global protein stability. It was shown that free cysteines on the surface of the protein globule which are not involved in the formation of inner disulfide bonds contribute to the insolubility of chitinase from Drosera capensis. The functional characteristics showed that chitinase exhibits high activity against colloidal chitin (360 units/g) and high fungicidal properties of recombinant chitinases against Parastagonospora nodorum. Latter highlights the application of chitinase from D. capensis as a promising enzyme for the control of fungal pathogens in agriculture.

最近，几丁质酶的研究已成为众多研究项目的重要目标，因为它们具有应用潜力，例如生物防治害虫剂。来自该属食肉植物的植物几丁质酶茅膏菜对多种植物病原体最具攻击性。然而，目标蛋白的低溶解性或不溶解性阻碍了几丁质酶作为生物杀真菌剂的应用。从该属的食肉植物中获得植物几丁质酶茅膏菜 以可溶形式在 大肠杆菌对于表达菌株，测试了三种不同的方法，包括透析、快速稀释和在 Ni-NTA 琼脂糖上复性复性。开发的“快速稀释”方案采用复性缓冲液补充 10% 甘油和 2M 精氨酸，结合还原/氧化谷胱甘肽的氧化还原对，将活性可溶性蛋白的产量增加到每 1 g 湿生物质 9.5 mg。为了提高几丁质酶的可溶性，基于结构的同源性建模，进行了基于结构的核心域中游离半胱氨酸的去除。鉴定了一种改进的几丁质酶变体 (C191A/C231S/C286T)，其在大肠杆菌表达系统与野生型相比。对野生型和改良变体的计算分析显示，在保持整体蛋白质稳定性的同时，结构的整体波动更大。结果表明，蛋白质球表面的游离半胱氨酸不参与内部二硫键的形成，导致几丁质酶的不溶性茅膏菜. 功能特性表明，几丁质酶对胶体几丁质具有高活性（360单位/g），重组几丁质酶对胶体几丁质具有高杀菌性能。结节拟孢霉。 后者重点介绍几丁质酶的应用 鸭嘴豆 作为一种有前途的酶，用于控制农业中的真菌病原体。