通过对大肠杆菌Escherichia coli的定向进化改善甜蛋白thaumatin II的可溶性表达
Lefei Wang1, Zilong Liu1, Liling Zhang2
1College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing 211816, China.
Journal of agricultural and food chemistry
|June 3, 2025
概括
我们使用工程Escherichia coli增强了天然甜蛋白thaumatin的表达. 定向进化产生了一种突变物,其蛋白质产量提高了45%,可溶性提高,并保留了用于工业应用的甜味.
科学领域:
- 生物技术是生物技术.
- 蛋白质工程是指蛋白质工程.
- 食品科学 食品科学 食品科学
背景情况:
- 陶马丁是一种天然的低热量甜蛋白,具有理想的特性,但由于溶解性有限和产量低,在微生物生产中面临挑战.
- 陶马丁的高效生产对于其作为食品工业中的天然甜味剂的应用至关重要.
研究的目的:
- 为了改造大肠杆菌以改善thaumatin II的可溶性表达.
- 通过蛋白质工程和定向进化来增强陶马丁II的溶解性和产量.
- 为了确认M-882突变的结构完整性,溶解性和甜味受体结合能力.
主要方法:
- 针对陶马丁II表达的工程大肠杆菌.
- 实现与分子伴侣一起表达,以提高溶解度.
- 应用定向进化来产生突变,随后进行结构和分子动力学分析.
- 评估了蛋白质产量,溶解度和甜味受体的结合亲和力.
主要成果:
- 与分子伴侣的同时表达显著改善了可溶性陶马丁II的表达.
- 定向进化产生了M-882突变,陶马丁产量增加了45% (42 mg/L).
- 突变M-882由于破坏二硫化物键,提高溶剂可访问性和增加结构灵活性,显示出更好的溶解性.
- 分子模拟和结合分析证实,M-882变体保留了甜味和受体结合能力.
结论:
- 与陪伴者共同表达和定向进化是增强thaumatin生产的有效策略.
- M-882陶马丁突变体在保持甜度的同时显示出更好的溶解性和产量,使其适合工业应用.
- 这项研究为为各种工业用途设计其他甜蛋白提供了框架.
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