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设计一种热稳定的迷你蛋白质,用于对重组蛋白和的蛋白质中介净化

Andrey A Karanov^1,2, Evgeniy A Zayats³, Maria A Kostromina³

¹Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, 117997, Russia. andrey-karanov2000@mail.ru.

Biochemistry. Biokhimiia

|July 3, 2025

概括

研究人员设计了一种来自*Thermus thermophilus* (TthDnaE1) 的热稳定迷你蛋白,该蛋白在高温下激活. 这种温度激活的整蛋白促进了高效的蛋白质拼接和裂变,用于重组蛋白质的生产.

科学领域:

生物化学生物化学
分子生物学分子生物学
蛋白质工程是指蛋白质工程.

背景情况:

整因是蛋白质拼接元件,可用于生物技术应用,可被设计成微型整因.
在需要高温的应用中,如重组蛋白质净化等,热稳定性质素具有优势.
通过温度激活来控制蛋白质活性对于精确的蛋白质操纵至关重要.

研究的目的:

为了设计和表征基于*Thermus thermophilus* DnaE1 (TthDnaE1) 的热稳定,温度激活的迷你体.
设计一种能够在高温下高效地进行蛋白质拼接或亲和标签的自我切割的微型蛋白质.
开发一种多功能工具,用于制造具有药学意义的重组蛋白和.

主要方法:

理性设计涉及删除突变 (Δ272, Δ280, Δ287) 在全长TthDnaE1.1.
单点突变发生 (C1A,D405G,C1A/D405G) 用于抑制外蛋白分裂和结合.
在不同温度下评估蛋白质拼接和裂变效率,最好在60°C左右.

主要成果:

两种微型蛋白质 (Δ272和Δ280) 在50°C以上表现出高效的蛋白质拼接.
Δ280 迷你引擎进一步进行了设计,以创建一个自切割的亲和度标签载体.
经过工程设计的TthDnaE1 Δ280 C1A/D405G小型蛋白在~60°C时显示出高的C端蛋白切割效率.

结论:

使用理性设计和TthDnaE1.1.的突变生成,成功构建了热稳定,温度激活的迷你体.
工程微型蛋白质提供了一个可控制的系统,用于蛋白质拼接和C端外蛋白分裂.
这种经过工程设计的TthDnaE1 Δ280 C1A/D405G微型蛋白质是开发用于制药蛋白质生产的新型表达系统的有希望的平台.

关键词:

结核酶域是一个内核酶域.迷你地铁中的迷你地铁再组合蛋白质和类.热稳定性热稳定性

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